\documentclass[10pt]{article}

\usepackage{fullpage}
\usepackage{setspace}
\usepackage{parskip}
\usepackage{titlesec}
\usepackage[section]{placeins}
\usepackage{xcolor}
\usepackage{breakcites}
\usepackage{lineno}
\usepackage{hyphenat}





\PassOptionsToPackage{hyphens}{url}
\usepackage[colorlinks = true,
            linkcolor = blue,
            urlcolor  = blue,
            citecolor = blue,
            anchorcolor = blue]{hyperref}
\usepackage{etoolbox}
\makeatletter
\patchcmd\@combinedblfloats{\box\@outputbox}{\unvbox\@outputbox}{}{%
  \errmessage{\noexpand\@combinedblfloats could not be patched}%
}%
\makeatother


\usepackage{natbib}




\renewenvironment{abstract}
  {{\bfseries\noindent{\abstractname}\par\nobreak}\footnotesize}
  {\bigskip}

\titlespacing{\section}{0pt}{*3}{*1}
\titlespacing{\subsection}{0pt}{*2}{*0.5}
\titlespacing{\subsubsection}{0pt}{*1.5}{0pt}


\usepackage{authblk}


\usepackage{graphicx}
\usepackage[space]{grffile}
\usepackage{latexsym}
\usepackage{textcomp}
\usepackage{longtable}
\usepackage{tabulary}
\usepackage{booktabs,array,multirow}
\usepackage{amsfonts,amsmath,amssymb}
\providecommand\citet{\cite}
\providecommand\citep{\cite}
\providecommand\citealt{\cite}
% You can conditionalize code for latexml or normal latex using this.
\newif\iflatexml\latexmlfalse
\providecommand{\tightlist}{\setlength{\itemsep}{0pt}\setlength{\parskip}{0pt}}%

\AtBeginDocument{\DeclareGraphicsExtensions{.pdf,.PDF,.eps,.EPS,.png,.PNG,.tif,.TIF,.jpg,.JPG,.jpeg,.JPEG}}

\usepackage[utf8]{inputenc}
\usepackage[greek,ngerman,english]{babel}



\usepackage{float}








\begin{document}

\title{Identification of patients with Fabry disease using routine pathology
results: PATHFINDER (eGFR) study}



\author[1]{Tim Reynolds}%
\author[2]{Karen Tylee}%
\author[2]{Kathryn Booth}%
\author[3]{Anthony Wierzbicki}%
\affil[1]{Burton Hospitals NHS Foundation Trust}%
\affil[2]{Manchester University NHS Foundation Trust}%
\affil[3]{Guy's and Saint Thomas' NHS Foundation Trust}%


\vspace{-1em}



  
  \date{May 04, 2024}


\begingroup
\let\center\flushleft
\let\endcenter\endflushleft
\maketitle
\endgroup





\selectlanguage{english}
\begin{abstract}
Aims: Lysosomal \selectlanguage{greek}α-\selectlanguage{english}galactosidase A deficiency (Fabry disease (FD)) was
considered an X-linked recessive disorder but is now viewed as a
variable penetrance dominant trait. The prevalence of FD is 1 in
40000-117,000 but the exact frequency is disputed depending on
ascertainment of late-onset cases and degree of female penetrance. Its
prevalence in the general population, especially in patients with
abnormal renal function is unclear. This study attempted to identify the
prevalence of FD in patients with abnormal results identified from
laboratory databases. Methods: Electronic laboratory databases were
interrogated to identify from clinical biochemistry records patients
with a phenotype of reduced estimated glomerular filtration rate
categorised by age on one occasion or more over a 3-year time interval.
Patients were recalled and a dried blood spot sample was collected for
determination of \selectlanguage{greek}α-\selectlanguage{english}galactosidase A activity by fluorimetric enzyme assay
in men and mass spectrometry assays of \selectlanguage{greek}α-\selectlanguage{english}galactosidase A and
lyso-globotriaosylceramide (lyso-GL-3) concentrations in women. Results:
Samples were obtained from 1084 patients identified with reduced renal
function. No cases of FD were identified in 505 men. From 579 women one
subject with reduced \selectlanguage{greek}α-\selectlanguage{english}galactosidase activity (1.5 \selectlanguage{greek}µ\selectlanguage{english}mol/l/hr) and
increased Lyso-GL-3 (5.5 ng/ml) was identified and shown to be
heterozygous for a FD mutation (c.898C\textgreater{}T; p.L300F;
Leu300Phe). It was later confirmed she was a relative of a known
affected patient. Conclusions: Pathology databases hold routine
information that can be used to identify patients with inherited errors
of metabolism. Biochemical screening using reduced eGFR has a low yield
for unidentified cases of Fabry Disease.%
\end{abstract}%



\sloppy




\emph{What is known:}

\begin{itemize}
\tightlist
\item
  Fabry's Disease is a treatable rare multisystem disorder affecting
  renal function with complex inheritance.
\item
  Its prevalence in the general population is unclear.
\end{itemize}

\emph{What this article adds:}

Screening laboratory databases using age-adjusted renal function
measures can identify patients with Fabry's disease but at a low yield

\emph{Introduction}

Lysosomal \selectlanguage{greek}α-\selectlanguage{english}galactosidase A deficiency causes Fabry disease
(FD)\textsuperscript{1-3}, which was considered to be X-linked recessive
and therefore females were simply carriers but more recently, it has
been shown to be a variable penetrance dominant trait as up to 70\%
heterozygous females are symptomatic\textsuperscript{4,5}. The
prevalence of Fabry's disease is 1 in 40000-117,000 but the exact
frequency is disputed depending on ascertainment of late-onset cases,
female penetrance and variants of unknown
significance\textsuperscript{6,7}. Deficiency of lysosomal
\selectlanguage{greek}α-\selectlanguage{english}galactosidase A activity causes accumulation of globotriaosylceramide
(Gb3), globotriaosylsphingosine (Lyso Gb3) and related
glycosphingolipids throughout the body\textsuperscript{8}. Clinical
symptoms are variable depending on age of onset and include abdominal
pain in 60\%, neuropathic pain and sensorineural deafness, multiple
angiokeratomas, renal failure, cardiomyopathy and
stroke\textsuperscript{1-3}.

Identification of patients with rare diseases is difficult. Case-finding
depends on the prior possibility of disease. Populations with end stage
renal disease (ESRD) on dialysis show a 10-fold Fabry Disease enrichment
compared to the general population\textsuperscript{7,9} and cases are
found in cardiology and stroke clinics\textsuperscript{7} but
unidentified cases may be difficult to locate in the wider population.
As enzyme replacement is now available, it would preferable to identify
patients prior to onset of renal failure\textsuperscript{10}. The
PATHFINDER project is based on the premise of using routine laboratory
results to identify patients who should be tested for inherited errors
of metabolism. The project design has been validated in patients with
other disorders\textsuperscript{11}. This paper describes the use of age
and creatinine-based estimated glomerular filtration rate (eGFR) as
screening biomarkers for FD.

\emph{Methods}

\textbf{Subjects}

Subjects were identified at 9 hospitals serving approx. 4,575,000 people
by interrogating laboratory databases to identify individuals in three
specific age bands with reduced eGFR calculated using the Modified Diet
in Renal Disease (MDRD; 4 variable) or chronic kidney disease
epidemiology (CKDEPI) equations. Thresholds were chosen empirically by
estimating a feasible population size that could be sampled based on the
distribution of eGFR results in a sample of data from Queen's Hospital,
Burton.

\begin{itemize}
\tightlist
\item
  Age 20-49, eGFR 15-54 mL/min/1.83m\textsuperscript{2}
\item
  Age 50-59, eGFR 15-49 mL/min/1.83m\textsuperscript{2}
\item
  Age 60-69, eGFR 15-39 ml/min/1.83m\textsuperscript{2}
\end{itemize}

Laboratory database searches were performed using creatinine
measurements over a maximum a 6-year time period interval limited by
site-specific data access issues. Once provisionally identified,
subjects were invited to take part in the study as they had been
identified as having abnormal results in routine pathology testing. They
were asked if they would meet with a research nurse to allow collection
of a blood sample for lysosomal \selectlanguage{greek}α-\selectlanguage{english}galactosidase activity. At the
research visit, patients provided informed consent, and basic
demographic details were collected. An EDTA blood sample was collected
and single drops were spotted on a dried blood spot (DBS) collection
card. Subjects indicated their preference for anonymity or if they
wanted to receive a copy of their results.

\selectlanguage{greek}\textbf{Lysosomal acid α-galactosidase analysis}\selectlanguage{english}

\textbf{Male samples}

Analysis of DBS cards was performed at a regional inherited errors of
metabolism laboratory. A 3 mm diameter disc was punched from the DBS and
extracted in sodium taurocholate by shaking at 4\selectlanguage{ngerman}°C for 60 min. Sample
extract was transferred to black 96-well microplates and incubated with
substrate for 20 hours at 37°C. Samples were analysed for
\selectlanguage{greek}α-\selectlanguage{english}galactosidase, and acarbose-inhibited acid \selectlanguage{greek}α-\selectlanguage{english}1,4-glucosidase, total
acid \selectlanguage{greek}α-\selectlanguage{english}1,4-glucosidase (measured as a reference enzyme) activities
measured simultaneously in different wells on the same
plate\textsuperscript{12}. The enzymatic reactions were stopped by the
addition of 100 \selectlanguage{greek}μ\selectlanguage{english}L/well sodium carbonate buffer (pH 9.5). Assays were
performed in duplicate, and one blank was assayed for each sample. To
prepare blanks, extract from each sample was added to incubated
substrate solution after the enzymatic reactions were stopped. The
relative fluorescence (excitation 360 nm, emission 460 nm) of each well
was measured using a Biotek Synergy HT fluorescence plate reader.
Fluorescence readings were corrected for the blank and compared with a
4- calibrator (supplied from Sigma)

The absolute amount of whole blood per spot is not known accurately but
is comparable between samples, so enzymatic activities are expressed as
pmol of substrate hydrolysed per punch per hour (pmol/punch/hour).
\selectlanguage{greek}α-\selectlanguage{english}galactosidase A activity below the reference range for males (6.3 --
47.0 pmol /punch/ hr) was considered significantly reduced. If activity
in the first sample was low, patients were invited to return for repeat
sampling and a fresh whole blood EDTA sample was sent for analysis.
Repeat samples were analysed in duplicate for alpha galactosidase
activity in leucocytes and plasma with beta-galactosidase and
beta-hexosaminidase (A and B) analysed as reference enzymes
respectively. Repeat assessment of blood spot activities of \selectlanguage{greek}α
-\selectlanguage{english}galactosidase and \selectlanguage{greek}α -\selectlanguage{english}1,4-glucosidase was also performed. Reference
ranges had been established by the local laboratory and had been
validated across UK specialist laboratories. Results were collated by
Queen's Hospital, Burton and communicated to participating sites.

\textbf{Female Samples}

Fabry disease is X-linked but females can still be affected. There is
overlap at the lower end of the \selectlanguage{greek}α-\selectlanguage{english}galactosidase A activity reference
range between affected and unaffected cases so a biomarker is also used.
Samples from females only were sent to ARCHIMEDlife laboratories
(Vienna, Austria) for assay of \selectlanguage{greek}α-\selectlanguage{english}galactosidase A activity and
Lyso-globotriaosylceramide (lyso-GL-3), a biomarker that is increased in
Fabry disease, by Tandem mass spectrometry\textsuperscript{8}. Results
from the female samples showing either a low enzyme activity, increased
lyso-GL-3 or both resulted in genetic sequence testing. \selectlanguage{greek}α-\selectlanguage{english}galactosidase
activity below the reference interval for females (1.2 -- 50.0 \selectlanguage{greek}µ\selectlanguage{english}mol/l
/hr) or Lyso-GL-3 above the reference limit (0-3.5ng/ml) were considered
significant. (comment: ref interval for a gal enzyme activity analysed
for the female samples at Archimed changed about the start of June 2019
to 2.8 - 50.0 umol/l/hr, as per letter issued, so if there are any
samples that were sent for analysis after this timeframe the new range
will have been used).

\emph{Results}

Database searches identified 1109 potential patients, with results
available for 1105. Protocol deviation was found in 21 cases with 3
subjects aged below 20yrs, and 18 over 70yrs leaving 1084 subjects in
total. Demographic information is shown in table 1.

\emph{Table 1 here}

In males \selectlanguage{greek}α-\selectlanguage{english}galactosidase A activity was 15.0\selectlanguage{ngerman}±5.23 (range 3.2-36.6)
pmol/punch/hr. Reduced \selectlanguage{greek}α-\selectlanguage{english}galactosidase activity
(\textless{}6.3pmol/punch/hr) was found in 7 male subjects from 505
tested. Low activity in DBS samples can be due to many factors including
a low white blood count, or inadequate sample drying. These subjects
were recalled for a second analysis: 2 subjects declined and 5 repeat
tests were normal. In females \selectlanguage{greek}α-\selectlanguage{english}galactosidase activity was 5.2\selectlanguage{ngerman}±2.4
(range 0.7-28.8) \selectlanguage{greek}µ\selectlanguage{english}mol/l/hr and 1 subject from 579 tested had reduced
\selectlanguage{greek}α-\selectlanguage{english}galactosidase activity (\textless{}1.2 \selectlanguage{greek}µ\selectlanguage{english}mol/l/hr). No pathogenic
mutations were found in patients with low \selectlanguage{greek}α-\selectlanguage{english}galactosidase activity. A
further 16 samples from females with normal range \selectlanguage{greek}α-\selectlanguage{english}galactosidase
activity had elevated Lyso-GL-3. One female subject (age 55, eGFR
40ml/min/1.73m\textsuperscript{2}) had \selectlanguage{greek}α-\selectlanguage{english}galactosidase activity 1.5
\selectlanguage{greek}µ\selectlanguage{english}mol/l/hr, but Lyso-GL-3 5.5 ng/ml and was identified as heterozygous
for a FD mutation (c.898C\textgreater{}T; p.L300F; Leu300Phe). She knew
that her mother had Fabry disease but had not been previously tested.

\emph{Discussion}

The prevalence of FD has been reported to be 1 in 40,000
-117,000\textsuperscript{6} but some sources suggest that as many as 1
in 50,000 males are affected and when late-onset disease is included the
incidence may be as high as 1 in 4600\textsuperscript{13,14}. The
prevalence of FD in patients with ESRD is 0.12-0.70\%. Earlier detection
would allow for treatment prior to the development of renal disease.
Neonatal blood spot screening for Fabry's is available in some US states
and Japan\textsuperscript{15}. This suggest a prevalence of 1 in 12,000
(8,000 if variants of unknown significance considered pathogenic).
Screening for FD does not form part of UK neonatal screening protocols.

Most case detection studies for FD have been conducted in patients with
significant renal disease or cardiomyopathy. Screening for Fabry's
disease in patients with CKD3b-4 from
0.2-0.6\%\textsuperscript{16-18}though some studies fail to find any
cases\textsuperscript{19}. A study of the prevalence of FD in a
secondary care population under the care of renal specialists is
underway in Australia\textsuperscript{6}. However, data on the
prevalence in earlier stages of renal disease (not under the care of
specialist renal physicians) is lacking. This study found one new female
case of FD from 1084 patients (505 male; 579 female) from primary care
and non-specialist secondary care based on age-related mild-severe renal
disease diagnosed on reduced eGFR. However \selectlanguage{greek}α-\selectlanguage{english}galactosidase activity for
females with FD overlaps with the bottom of the normal range resulting
in up to 30\% false negatives so addition of lysoGL-3 is recommended in
females\textsuperscript{20,21}. Enhanced screening with LysoGL-3
biomarker confirmed one female was heterozygous for FD
(c.898C\textgreater{}T; p.L300F ; Leu300Phe)\textsuperscript{22}.
Further work identified that she was related to a known affected case
but had not been contacted for family screening.

The PATHFINDER study pathology-based electronic health record data to
identify patients from primary care and non-specialist secondary care
mostly CKD stage 2-3(a). This limited this study to use only creatinine
(eGFR) data rather than complete renal disease
profiles\textsuperscript{23}. No data on albuminuria or (less relevant)
haematuria. Albuminuria is considered a useful marker for FD but many
early cases of FD lack significant albuminuria\textsuperscript{24}. The
cohort sampled was lower risk than previous studies and included both
primary and non-specialist secondary care settings. Though underpowered
to detect significant numbers of cases, it suggests that enrichment for
prior renal disease using a CKD3 cut-off and possibly additional
biomarker stratification\textsuperscript{25} is required before
screening for FD is viable.

\emph{}

\emph{PATHFINDER Project Collaborator group:}\selectlanguage{english}
\begin{longtable}[]{@{}lllll@{}}
\toprule
\begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
Collaborating Centre\strut
\end{minipage} & \begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
Research team\strut
\end{minipage} & \begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
Creatinine assay and eGFR calculation method\strut
\end{minipage} & \begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
Approx population served\strut
\end{minipage} & \begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
No. of Recruits\strut
\end{minipage}\tabularnewline
\midrule
\endhead
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Ipswich Hospital\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Dr Taruna Likhari (PI), Genessa Peters (RN)\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Enzymatic creatinine CKD-EPI\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
385000\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
104\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Princess Royal Hospital, Telford\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Dr Nigel Capps (PI), Louise Tonks (RN)\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Enzymatic creatinine MDRD until 30/10/18, then CKD-EPI\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
490000\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
104\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Queen's Hospital, Burton\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Professor Tim Reynolds (PI), Louise Wilcox (RN), Clare Mewies (SA)\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Jaffe creatinine CKD-EPI\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
360000\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
68\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Russell's Hall Hospital, Dudley\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Ms Jackie Smith (PI \& RN)\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Enzymatic creatinine CKD-EPI\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
450000\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
430\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Oxford University Hospital\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Mrs Nicky McRobert (PI), Jamie Burbage (RN)\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Enzymatic creatinine CKD-EPI\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
655000\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
85\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
University Hospital North Midlands\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Dr Anthony Fryer (PI), Loretta Barnett, Susan Hendy (RN)\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Enzymatic creatinine CKD-EPI\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
900000\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
50\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Royal Berkshire Hospital\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Julie Sutton (PI \& RN)\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Enzymatic creatinine CKD-EPI\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
600000\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
162\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Brighton \& Sussex University Hospitals\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Prof Gordon Ferns (PI) , Mel Smith, Valentina Toska (RN)\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Jaffe creatinine MDRD\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
460000\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
40\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
West Suffolk Hospital\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Prof Patrick Twomey (PI) Joanne Kellett (RN)\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Enzymatic creatinine CKD-EPI\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
275000\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
62\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
Total number of recruits\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
Total number of recruits\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
Total number of recruits\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
4,575,000\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
1105\strut
\end{minipage}\tabularnewline
\bottomrule
\end{longtable}

Population served extracted from individual hospital CQC reports
Abbreviations: Principal Investigator (PI); RN = Research Nurse (RN);
Study administrator (SA)

\textbf{Contributors} : The study was designed by TMR, who also ran the
study data gathering and management of the sample and results.
Specialist biochemical and genetic analyses were performed by KT and KB.
Data were analysed by TMR and ASW, and they wrote the study manuscript.
The final manuscript as submitted was approved by all the authors.

\textbf{Funding} : This work was supported by an investigator grant from
Sanofi Genzyme Therapeutics, Oxford. Research nurses were funded by the
National Institute for Health Research.

\textbf{Competing interests} : TMR is currently in receipt of project
grants from Genzyme Therapeutics, Oxford, UK (now Sanofi Genzyme,
Oxford, UK); Shire Pharmaceuticals, Basingstoke, UK, now Takeda
Pharmaceutical Ltd; and Synageva BioPharma, Watford, UK (now Alexion
Pharma UK, Uxbridge, UK).

\textbf{Patient consent for publication} : Not required.

\textbf{Ethics approval} : The project received ethics approval from the
National Research Ethics Service Committee East Midlands - Northampton
(UK Integrated Research Application System project number 158121;
14/EM/1153; 14/10/2014). It was included in the National Institute for
Health Research portfolio (UKCRN ID: 17588) and transferred to Health
Research Authority on 15 June 2016.

\emph{References}

1. Hsu TR, Niu DM. Fabry disease: Review and experience during newborn
screening. \emph{Trends Cardiovasc Med.} 2018;28(4):274-281.

2. Schiffmann R. Fabry disease. \emph{Handb Clin
Neurol.}2015;132:231-248.

3. Platt FM, d'Azzo A, Davidson BL, Neufeld EF, Tifft CJ. Lysosomal
storage diseases. \emph{Nat Rev Dis Primers.} 2018;4(1):27.

4. Wang RY, Lelis A, Mirocha J, Wilcox WR. Heterozygous Fabry women are
not just carriers, but have a significant burden of disease and impaired
quality of life. \emph{Genet Med.} 2007;9(1):34-45.

5. Deegan PB, Bahner F, Barba M, Hughes DA, Beck M. Fabry disease in
females: clinical characteristics and effects of enzyme replacement
therapy. In: Mehta A, Beck M, Sunder-Plassmann G, eds. \emph{Fabry
Disease: Perspectives from 5 Years of FOS.} Oxford2006.

6. Mallett A, Kearey P, Cameron A, et al. The Ckd. Qld fabRy
Epidemiology (aCQuiRE) study protocol: identifying the prevalence of
Fabry disease amongst patients with kidney disease in Queensland,
Australia. \emph{BMC Nephrol.} 2020;21(1):58.

7. Doheny D, Srinivasan R, Pagant S, Chen B, Yasuda M, Desnick RJ. Fabry
Disease: prevalence of affected males and heterozygotes with pathogenic
GLA mutations identified by screening renal, cardiac and stroke clinics,
1995-2017. \emph{J Med Genet.} 2018;55(4):261-268.

8. Aerts JM, Groener JE, Kuiper S, et al. Elevated
globotriaosylsphingosine is a hallmark of Fabry disease. \emph{Proc Natl
Acad Sci U S A.} 2008;105(8):2812-2817.

9. Thadhani R, Wolf M, West ML, et al. Patients with Fabry disease on
dialysis in the United States. \emph{Kidney Int.} 2002;61(1):249-255.

10. Ortiz A, Germain DP, Desnick RJ, et al. Fabry disease revisited:
Management and treatment recommendations for adult patients. \emph{Mol
Genet Metab.} 2018;123(4):416-427.

11. Reynolds TM, Mewies C, Hamilton J, Wierzbicki AS, group PPC.
Identification of rare diseases by screening a population selected on
the basis of routine pathology results-the PATHFINDER project: lysosomal
acid lipase/cholesteryl ester storage disease substudy. \emph{J Clin
Pathol.} 2018;71(7):608-613.

12. Chamoles NA, Niizawa G, Blanco M, Gaggioli D, Casentini C. Glycogen
storage disease type II: enzymatic screening in dried blood spots on
filter paper. \emph{Clin Chim Acta.} 2004;347(1-2):97-102.

13. MacDermot KD, Holmes A, Miners AH. Natural history of Fabry disease
in affected males and obligate carrier females. \emph{J Inherit Metab
Dis.} 2001;24 Suppl 2:13-14; discussion 11-12.

14. Spada M, Pagliardini S, Yasuda M, et al. High incidence of
later-onset fabry disease revealed by newborn screening. \emph{Am J Hum
Genet.} 2006;79(1):31-40.

15. Sawada T, Kido J, Yoshida S, et al. Newborn screening for Fabry
disease in the western region of Japan. \emph{Mol Genet Metab
Rep.}2020;22:100562.

16. Nakao S, Kodama C, Takenaka T, et al. Fabry disease: detection of
undiagnosed hemodialysis patients and identification of a ''renal
variant'' phenotype. \emph{Kidney Int.} 2003;64(3):801-807.

17. Mehta A, Beck M, Eyskens F, et al. Fabry disease: a review of
current management strategies. \emph{QJM.} 2010;103(9):641-659.

18. Capuano I, Garofalo C, Buonanno P, et al. Identifying Fabry patients
in dialysis population: prevalence of GLA mutations by renal clinic
screening, 1995-2019. \emph{J Nephrol.} 2019.

19. Jahan S, Sarathchandran S, Akhter S, et al. Prevalence of Fabry
disease in dialysis patients: Western Australia Fabry disease screening
study - the FoRWARD study. \emph{Orphanet J Rare Dis.} 2020;15(1):10.

20. Linthorst GE, Vedder AC, Aerts JM, Hollak CE. Screening for Fabry
disease using whole blood spots fails to identify one-third of female
carriers. \emph{Clin Chim Acta.} 2005;353(1-2):201-203.

21. Stiles AR, Zhang H, Dai J, et al. A comprehensive testing algorithm
for the diagnosis of Fabry disease in males and females. \emph{Mol Genet
Metab.} 2020.

22. Shabbeer J, Robinson M, Desnick RJ. Detection of alpha-galactosidase
a mutations causing Fabry disease by denaturing high performance liquid
chromatography. \emph{Hum Mutat.} 2005;25(3):299-305.

23. National Institute for Health and Care Excellence. Chronic kidney
disease in adults: assessment and management. Clinical guideline
(CG182). 2014; NICE CKD. Available at:
https://www.nice.org.uk/guidance/cg182. Accessed 1/4, 2020.

24. Riccio E, Sabbatini M, Capuano I, Pisani A. Early Biomarkers of
Fabry Nephropathy: A Review of the Literature.
\emph{Nephron.}2019;143(4):274-281.

25. Voorink-Moret M, Goorden SMI, van Kuilenburg ABP, et al. Rapid
screening for lipid storage disorders using biochemical markers. Expert
center data and review of the literature. \emph{Mol Genet
Metab.}2018;123(2):76-84.

\textbf{Hosted file}

\verb`IJCP Pathfinder Fabry table.docx`  available at \url{https://authorea.com/users/331989/articles/458478-identification-of-patients-with-fabry-disease-using-routine-pathology-results-pathfinder-egfr-study}

\selectlanguage{english}
\FloatBarrier
\end{document}

