Hematopoietic stem cell transplantation for purine nucleoside phosphorylase deficiency: an EBMT-IEWP retrospective study

Key Points

• •HSCT yields satisfactory long-term outcomes in patients with PNP deficiency.
• •OS was best in early-diagnosed patients without neurologic symptoms.

Visual Abstract

Abstract

Purine nucleoside phosphorylase (PNP) deficiency causes inadequate purine metabolite detoxification, which leads to combined immunodeficiency and variable neurologic symptoms. Hematopoietic stem cell transplantation (HSCT) cures the immunodeficiency, but large studies on the long-term outcomes are lacking. In a retrospective study of the European Society for Blood and Marrow Transplantation, we investigated 46 patients with PNP deficiency from 21 centers. We analyzed the presenting clinical signs and outcomes after HSCT. Cognition (0-3), hearing (0-3), interaction (0-4), movement (0-4), and occupation (0-3) (CHIMO) were scored at the last follow-up (FU) visit (no impairment, 17; mild, 15-16; moderate, 12-14; and severe impairment, <12). The median age at initial presentation was 7.5 (1-48) months. The patients presented with infections (41%), neurological dysfunction (39%), both (15%), or autoimmune disease (5%). At the time of HSCT (median age, 26 [2-192] months), neurological abnormalities were observed in 88% of patients. After a median FU of 7.9 (1.0-22.3) years, 40 patients were alive with a 3-year overall survival (OS)/event-free survival (EFS) probabilities of 86% (confidence interval [CI], 77%-97%)/75% (CI, 64%-89%), respectively. High-level (>50%-100%)/low-level donor chimerism (11%-50%) was observed in 85%/15% of patients, respectively, leading to resolution of T lymphopenia. The median overall CHIMO score was 14 (6-17), while the median scores for each component were 3 (0-3), 3 (1-3), 4 (1-4), 3 (1-4), and 2 (0-3), respectively. Patients who underwent HSCT before 24 months after the initial presentation demonstrated superior OS (P = .049). Neurological symptoms that occurred before 11 months of age were associated with reduced OS (P = .027). While the overall results were satisfactory, earlier diagnosis could further improve outcomes.

Purine nucleoside phosphorylase (PNP) deficiency is a rare inborn error of immunity manifesting as progressive loss of T-cell number and function, which may be preceded by progressive neurological dysfunction. In a retrospective study, Herrmann et al report that timely hematopoietic stem cell transplantation can efficiently correct the immune defects caused by PNP deficiency and stabilize the neurologic abnormalities. These data provide a benchmark for the management of this devastating disease.

Introduction

Purine nucleoside phosphorylase (PNP) deficiency is an autosomal recessive inherited disorder of purine metabolism that affects ∼1% to 2% of all patients with combined immunodeficiency (CID).^{1, 2, 3, 4, 5, 6, 7, 8} PNP is an enzyme that reversibly catalyzes the phosphorolysis of inosine, deoxyinosine, guanosine, and deoxyguanosine^{1, 2, 3} Absent or significantly reduced PNP enzyme activity leads to increased levels of deoxyguanosine and deoxyinosine in plasma, cerebrospinal fluid, and urine.^1,4 The deoxytriphosphate compounds, especially deoxyguanosine triphosphate, accumulate intracellularly and induce apoptosis in lymphocytes and neuronal cells, and T lymphocytes are more susceptible to apoptosis than B lymphocytes.^4,5 The diagnosis can be confirmed by determining the purine levels in body fluids using high-performance liquid chromatography.^6,7 All individuals with biallelic pathogenic variants in PNP are symptomatic.⁸

It has been reported that clinical symptoms in PNP deficiency occur at a later stage than in classical severe CID (SCID), particularly between infancy and preschool age.¹ Newborn screening for SCID, based on the detection of low numbers or no cells that bear T-cell receptor excision circles, seems to detect only a minority of individuals with PNP deficiency.^{3,8, 9, 10, 11, 12, 13, 14, 15} The more sensitive tandem mass spectrometry analysis of purine metabolites using dried blood spots is unfortunately not routinely available.^10,16

Immunodeficiency in PNP deficiency is characterized by the occurrence of recurrent and/or severe bacterial, viral, or fungal infections,^17,18 as well other opportunistic infections.^11,19,20 In addition, there is a predisposition to autoimmune diseases (AID+), particularly autoimmune cytopenia, and less commonly to macrophage activation syndrome and hematologic malignancies.^15,21,22 Up to two-thirds of patients exhibit heterogeneous neurologic symptoms, including developmental delay and neurologic and cognitive deficits.^{1,23, 24, 25, 26, 27} Neurologic involvement varies clinically from normal to severely impaired, even within families with the same molecular defect.^17,18,24

Allogeneic hematopoietic stem cell transplantation (HSCT) is the only curative treatment.^{15,23,24,28,29} Transient metabolic improvement can be achieved with erythrocyte exchange transfusions (ET),^30,31 but unlike for adenosine deaminase deficiency, no enzyme replacement therapy is available.³² There are anecdotal reports that neurologic status improves in some patients after HSCT,^15,24,28 whereas neurologic damage persists in others;²³ thus, the benefit of HSCT on the long-term neurologic outcomes is largely unknown.

In this study, we provide a comprehensive analysis of the onset and nature of clinical presentation and clinical outcomes in a patient cohort of 46 children with PNP deficiency who underwent HSCT.

Patients, materials, and methods

Study design

In this multicenter, retrospective study of the Inborn Errors Working Party (IEWP) of the European Society for Blood and Marrow Transplantation (EBMT) (study number 8427029), centers in Europe, Canada, and the United States enrolled patients with proven PNP deficiency who underwent transplantation between 1998 and 2022. The criteria for inclusion required at least 1 of the following: (1) absent or severely deficient PNP enzyme activity (<5% of normal as measured in erythrocytes from untransfused patients) and (2) genetic confirmation of homozygous or compound heterozygous mutations known or predicted to cause PNP deficiency or already detected in a sibling affected with PNP deficiency. Related donors were tested for PNP mutations and PNP activity. Healthy family donors with heterozygous PNP mutations were permitted to donate. Participants or their legal guardians provided written informed consent to participate in this study via the EBMT for Europe and Primary Immune Deficiency Treatment Consortium (PIDTC) for Canada and the United States.

Data collection and definitions

Data were collected using specially designed case report forms and included information on the baseline disease-specific characteristics, including infectious, neurologic, and autoimmune symptoms, and laboratory data at clinical presentation and HSCT. The outcome analysis included information on conditioning chemotherapy,^{33, 34, 35} serotherapy, graft source and in vitro manipulation, post-HSCT immunosuppression, primary and secondary graft failure (GF), the need for a second HSCT or cellular therapy (CT; donor lymphocyte infusion [DLI], stem cell boost [SCB] infusion), incidence of acute and chronic graft-versus-host disease (GVHD), and infectious and autoimmune complications (supplemental Material and methods, available on the Blood website).

At the last follow-up visit (FU), data analysis included overall survival (OS; days alive after HSCT), event-free survival (EFS; days alive without a second HSCT or clinical relapse),³³ incidence of acute (Glucksberg score) and chronic GVHD (limited, extensive), the percentage of donor chimerism (DC) in the whole blood and/or in sorted myeloid (CD15⁺neutrophils/CD14⁺monocytes), T- (CD3⁺), and B-cell (CD19⁺) compartments, independence of immunoglobulin G replacement therapy, and, if available, PNP enzyme activity in blood and/or urinary purine metabolite excretion (uric acid, deoxyguanosine triphosphate, guanosine, inosine, deoxyguanosine, and deoxyinosine) according to established local laboratory methods.

At the final FU, the study physicians assessed the neurologic outcomes using a new combination of standardized and validated scores,^{36, 37, 38, 39, 40} including cognition (scoring range, 0-3), hearing (0-3), interaction (0-4), movement (0-4), and occupation (0-3), referred to as the CHIMO score (maximum: 17). Karnofsky and/or Lansky scores were also recorded (supplemental Tables 1 and 2).

Statistical analysis

Data on the clinical and laboratory characteristics were collected at the time of initial diagnosis, at HSCT, and at the last FU (at least 1 year after HSCT). Categorical variables were compared using Fisher exact test and continuous variables were compared using the Mann-Whitney U test. Correlations were calculated using Spearman's rank correlation coefficient. For survival, Kaplan-Meier estimates were used for survival and log-rank tests were used for comparison. Data were censored at the date of last FU for surviving patients. Confidence intervals (CIs) were calculated using a log-log transformation. Kaplan-Meier curve comparisons between groups were performed using the log-rank test. The Cox proportional hazards model was used for univariate analysis of the risk factors associated with EFS and OS. For the Cox proportional hazard regression and depiction, the coxph and ggforest functions were used in R, and to determine the optimal cutoff timepoints in the survival analysis, the surv_cutpoint function in the survminer package in R was applied. A multivariate analysis was not performed because of the small sample size. All analyses were performed using RStudio version 2024.09.0, packages dplyr, forestplot, ggthemes, knitr, survminer, survival, tidyquant, and tidyverse, and Microsoft Excel and Graphpad Prism V9 and 10.

Institutional review board approval was obtained for participation in the study on behalf of the EBMT IEWP.

Results

Patient characteristics

PNP deficiency was diagnosed at a median age of 17.5 months (0-186), including 6 asymptomatic patients (13%) who were diagnosed at birth based on family history. Another patient with a family history of CID was diagnosed at the onset of neurologic symptoms. Allogeneic HSCT was performed at a median age of 25.5 months (2-192). Five of the 6 perinatally diagnosed patients were transplanted before the onset of symptoms. Most patients fulfilled the criteria for leaky T cell-negative, B cell-positive, natural killer cell-positive severe combined immunodeficiency (T-B+NK+SCID) or CID⁴¹ (Table 2), and none had reported Omenn syndrome or the presence of maternal lymphocytes. Infectious symptoms without neurologic compromise (I+) were presenting symptoms in 41% (17/41) of patients, neurologic symptoms without signs of immunodeficiency (N+) were the presenting symptoms in 39% (16/41), and combined infectious and neurologic symptoms (I+/N+) were the presenting symptoms in 15% (6/41). AID+ was a presenting symptom in 5% (2/41) of patients, either with or without infectious symptoms. Patients with only 1 type of initial clinical manifestations (I+ or N+ or AID+) presented earlier than patients with a combined occurrence of different types of initial clinical manifestations. The median age at first presentation of I+, N+, AID+, and combined I+/N+ was 5, 8, 13.5, and 15 months, respectively (Table 1). The overall distribution of all N+, I+, and A+ symptoms over time is shown in Figure 1A.

Table 2.

Immunologic phenotype, pretransplantation erythrocyte PNP enzyme activity, and mutations of the PNP gene

Patient ID	CD3 T (per μL)	CD4/45RA T (per μL)	B (per μL)	NK (per μL)	ANC (per μL)	Eos (per μL)	IgG (g/L)	IgA (g/L)	IgA (g/L)	IgM (g/L)	Erythrocyte PNP activity∗	Mutation
1†	7	N.a.	33		1850	56	11	0.75	0.75	0.82	Undetectable
2	96	N.a.	61	233	4730	0	7.8	0.93	0.93	2.23	Undetectable
3	65	N.a.	24	250	1100	100	18.8	1.8	1.8	1.15	Undetectable
4†	336	N.a.	25	274	350	775	9.9	0.57	0.57	1.99	Undetectable
5	230	N.a.	32	750	3420	340	8.3	1.29	1.29	1.58	Undetectable
6	48	N.a.	30	43	3600	90	8.5	0.47	0.47	1.53	Undetectable
7	37	48	14	132	300	0	5.5	0.73	0.73	0.14	Undetectable	Homozygous intronic variant c286-18G>A
8	65	65	88	216	4800	100	9.8	1.07	1.07	1.08	Undetectable	Homozygous intronic variant c286-18G>A
9	444	444	48	60	6700	200	10.4	1.07	1.07	1.18	Undetectable	Nonsense c.172C>T (R58X)
10	28	N.a.	40	220	810	90	7.1	0.58	0.58	1.15	Undetectable
11	1131	N.a.	48	320	7128	648	1.88	0.71	0.71	0.5	Reduced
12	1160	N.a.	420	139	4510	896	786	0.95	0.95	0.85	Undetectable
13	13	N.a.	57	110	3200	150	24.5‡	<0.2	0.2	1.1	Undetectable
14	92	5	67	260	940	40	13.3§	0.53	0.53	1.36	Reduced
15	20	4	30	40	700	70	9.33	0.52	0.52	0.4	Undetectable
16ǁ	658	N.a.	102		212	64	0.3	N.a.	0.07	0.23	Undetectable
17	12	N.a.	9	52	3100	200	<0.3	0.04	0.04	<0.05	Reduced	Homozygous c.383A>G
18	348	N.a.	76	242	250	650	16.4	0.53	0.53	1,8	Undetectable
19	120	50	60	150	0	780	6.7	0.07	0.07	n/a	Undetectable
20	207	N.a.	100	205	5170	110	7.13	0.5	0.5	1.83	Reduced	Homozygous missense c.117A>T
21	145	N.a.	29	189	3700	400	3.42	0.09	0.09	0.42	Undetectable
22	59	N.a.	25	330	1190	275	3.4	0.06	0.06	0.04	Undetectable
23	99	6	26	39	2542	101	7.88	0.078	0.078	1.12	Reduced	Compound-heterozygous Exon 2 c.58CGA>TGA Exon 3 c.98GAA>TAA
24	172	38/0	37	159	2120	180	8.58	0.059	0.059	2.08	Reduced	Compound heterozygous Exon2 c.172C>T p.Arg58Stop Exon 4 c.389T>C p.Met130Thr
25†	130	3	150	170	1190	410	13.3	0.53	0.53	1.36	Undetectable
26ǁ	50	9	30		5180	370	n.d.		0.08	0.09	Reduced
27	70	N.a.	50	190	3687	209	12.2	0.9	0.9	1.7	Undetectable
28	117	N.a.	75	158	2003	940	5.9	0.98	0.98	0.75	Reduced
29†	80	47	53	118	484	66	0.2	0	0	4.2	Undetectable	Compound heterozygous Exon 2 Arg24Termc. 70 C>T Exon 3 Glu89. Lys c.265G>A
30	43	N.a.	24	156	5200	190	10	0.3	0.3	0.9	Undetectable
31	10	0	100	70	760	120	7.94	0.66	0.66	0.41	Reduced	Homozygous c.383A>G, p.D128G
32	250	10	57	157			11.5		0.71	0.38	Undetectable	c.487T>C, p.Ser163Pro
33	369	35	101	93	2700	7	1.67	0.04	0.04	0.18	Undetectable
34	19	0	15	26	6300	0.2	8.99	0.96	0.96	41	Reduced	Compound heterozygous c.234R>P; c.212L>P
35	170	15	161	55	6400	0.1	8.29	0.88	0.88	63	Not done	Compound heterozygous c.234R>P; c.212L>P
36†	158	2	72	531	4000	0.14	11.8	1.09	1.09	133	Undetectable	Compound heterozygous
37ǁ	1299	185	143		5150	176	11.85¶	N.a.	0.04	0.03	Reduced	Homozygous c.286-18G>A
38ǁ	511	239	23		9460	120	11.2¶	N.a.	0.04	0.05	Reduced	Homozygous c.286-18G>A
39	60	9	40	80	1400	200	8.82	0.75	0.75	1.85	Undetectable	c.172C>T, p(ARG58∗); c.701G>C, p.(Arg234Pro)
40	90	14	120	N.a.	3000	1600	10.35	N.a.	0.42	1.29	Undetectable	c.172C>T, p(ARG58∗); c.701G>C, p.(Arg234Pro)
41ǁ	2510	1711	320	N.a.	6200	400	2.05	N.a.	0.06	0.7	Undetectable	c.172C>T, p(ARG58∗); c.701G>C, p.(Arg234Pro)
42	34	22/0	165	290	3710	180	10.5	0.06	0.06	1.07	Undetectable	Homozygous c.59A>C(p.H20P)(p.His20Pro)
43	36	N.a.	88	372	1420	510	2.35	<0.22	<0.22	0.37	ND	Homozygous c.172C>T
44†	38	N.a.	3	59	22 400	0	9.56		0.43	0.33	Undetectable	Homozygous c.286-2A>T
45	170	19	30	160	5.49	0.04	1.1	<0,1	<0,1	1.28	Reduced	Homozygous c.265G>A, p.Glu89Lys
46ǁ	830	103	50	260	3.43	0.06	5.17	0.16	0.16	0.33	Undetectable	Homozygous c.244C>T, p.GLN82∗

Immunologic parameters at diagnosis.

ANC, absolute neutrophil count; Eos, eosinophils; ID, identifier; Ig, immunoglobulin; N.a., not available; NK, natural killer cells.

^∗Untransfused patients (according to local laboratory normal ranges and SI units).

^†Deceased patient.

^‡Monoclonal IgG kappa.

^§On intravenous immunoglobulin replacement therapy.

^ǁNeonatal diagnosis owing to family history (patients 37, 38, 41, and 46 with erythrocyte exchange transfusions after birth).

^¶Maternal IgG.

Table 1.

Clinical presentation

ID	Sex	Initial neurologic findings	Initial I+ and AID+ symptoms	Type of initial presentation	I+ present age, mo	N+ present age, mo	AID+ present age, mo	PNP diagnose age, mo	HSCT at age, mo
1∗	F	Tremor, ataxia, DL	RTI, GI (Giardia)	N	30	6		30	35
2	F	DL in motor skills	None, VZV (treated with acyclovir)	N	15	6		15	26
3	M	DL	RTI, OM, diarrhea	I	1	30		47	51
4∗	F	DL, microcephalus/brachycephalus, encephalitis/meningitis (at 2 y): subsequent hydrocephalus, ataxia, spastic paresis of upper limbs	Diarrhea RTI, OM	I	4	24		24	35
5	M	DL, generalized hypotonia	Diarrhea, VZV/pneumonia, OM	I	6	9		9	14
6	F	Ataxia neck/trunk, choreatic movements, polymicrogyria (like sibling not suffering from PNP)	RTI	N	6	3		6	10
7	M	DL, spastic paresis	Family history (brother had CID)	N		12		12	27
8	F	DL in motor skills, mild truncal hypotonia	Severe VZV	I	11	12		12	16
9	F	DL	RTI, GI, thrush	I	5	11		11	14
10	F	DL, spastic paraparesis, MRI: global atrophy, corpus callosum atrophy	RTI, VZV (pneumonia/myocarditis)	N	23	6		23	25
11	F	Hypotonia, insufficient head control	Severe VZV, septicemia	I	3	7		3	6
12	M	Motoric coordination disorder, spastic paresis	RTI, impetigo, GI (RV)	I	12	24		71	72
13	F	Ataxia (MRI: normal), after first HSCT ability to walk; deterioration before second HSCT, motoric DL	Thrush, RTI (RSV), GI (RV), AIHA	I	7	20	20	22	24
14	M	Lower limb/truncal hypotonia; spastic paresis; motor DL with impaired balance	OM, RTI, severe VZV, zoster	N	20	9		9	47
15	M	SNHL unrelated to PNP (other cause; a cochlear implant); mild gross motor DL, consanguinity	AIHA, ITP, URT, RTI/bronchiectasis; intussusception	A/I	23	None	23	23	39
16	F	Normal	Family history† (newborn diagnosis)	None		None		0	2
17	F	Generalized hypotonia, DL	RTI, OM; CMV (including CNS)	I/N	15	15		15	39
18	F	DL of motor and speech function	AIHA, ITP, pyelonephritis; sepsis	I/N	18	18		18	25
19	M	Spastic paresis, mild spastic dysplasia, DL	AIN, RTI	I/N	19	19	19	19	22
20	F	Cerebral palsy and spastic paraparesis, DL	RTI, VZV, bronchiectasis, thrush	N	72	8		72	168
21	M	Ataxia, spastic paraparesis, mild DL	RTI, thrush, dermatitis	I	1	18		26	28
22	F	Normal	Thrush, GI (Campylobacter) (sibling died from EBV/lymphoma)	I	24	None		24	25
23	M	DL, motor retardation with spastic paresis of lower extremities, progressed to spastic tetraparesis	RTI, OM	I/N	12	12		12	42
24	F	Normal	AIHA, ITP	A	6	None	6	6	12
25∗	M	Hemiplegia, gross motor DL; MRI brain: delayed myelination/thin corpus callosum; bilateral lower motor neuron signs in both lower limbs	RTI, paronychia, AIN, CMV/EBV	N	11	6	11	6	31
26	F	Upper motor signs at HSCT; motor DL present with asymmetry of tone increased in lower limbs and posturing of upper limbs/head	Family history: sibling had severe neurologic disease and died without HSCT (newborn diagnosis)	N		5		0	7
27	F	DL, atactic, unstable gait, motor DL; after severe VZV; central motor paralysis	AIHA; severe VZV	N	18	12	18	18	36
28	M	Normal	Severe VZV	I	36	None		38	40
29∗	M	Mild DL, hypotonia	Recurrent thrush	I	14	10		21	22
30	F	Severe DL with no sitting/crawling/running	RTI, tracheomalacia	N	12	8		12	20
31	F	Generalized hypotonia Ataxia (noted after general anesthesia). Unsafe swallow	RTI (influenza A). GI (NV, SP, RV)	I	4	16		4	18
32	M	Choreoathetosis, motor DL	VZV (at 48 mo of age), DLBCL (EBV-related at 186 mo of age)	N	48	20		186	192
33	M	Ataxia	Thrush	I	12	14		18	43
34	M	Stiffness both lower extremities and ankles; mild, nonprogressive gait difficulty	Atopic dermatitis asthma; sinusitis, vaccine related VZV (via sibling)	N	48	12		108	119
35	M	Motor DL, ataxia, spastic diplegia; MRI: normal	Asthma, cow's milk/peanut allergy, vaccine related VZV, RTI	N	18	18		84	92
36∗	M	Hypotonia, motor and speech delay	OM, thrush, zoster (VZV); CMV/ADV	N	48	9		48	52
37	F	Normal	Family history† (newborn diagnosis)	None		None		0	2
38‡	F	Normal	Family history† (newborn diagnosis)	None		None		0	4
39‡	M	Grossly normal examination; ataxia, hemi spastic paresis; motor delay	RTI, AIHA, dermatitis, zoster (VZV)	I	1	11	11	35	41
40‡	M	Motor DL, axial hypotonia	Family history (patient 39); cleft palate; OM with conductive hearing loss	I/N	12	12		17	26
41‡	M	Normal	Family history (sibling of patient 39/40) (antenatal diagnosis)†	None		None		0	4
42	M	Generalized hypotonia	RTI (ADV/CV) diarrhea, thrush	I	5	7		11	12
43	F	Motor DL, generalized hypotonia	Aphthous stomatitis, EBV-LGN (CNS, kidney, spleen, liver)	I	3	17		17	22
44∗	F	Spastic diplegia, DL	MAS and arthritis	N		5	31	33	37
45	F	Motor DL, spastic tendency, ataxia, hypotonia	Vaccine related VZV	I/N	21	6		21	23
46	F	Normal	Family history† (newborn diagnosis)	None		None		0	3

ADV, adenovirus; CV, coronavirus; DL, developmental delay; DLBCL, diffuse large B-cell lymphoma; LGN, lymphoid granulomatosis; GI, gastrointestinal infection; I+, immunodeficiency; MAS, macrophage activation syndrome; N+, neurologic abnormalities; OM, otitis media; NV, norovirus; RV, rotavirus; RTI, respiratory tract infection; SP, sapovirus; URT, urinary tract infection.

^∗Deceased patient.

^†Erythrocyte exchange transfusion after birth.

^‡Siblings of 1 family.

Figure 1.

Age and clinical manifestations of the cohort. (A) Distribution of the age at first symptom onset (x-axis) across different initial clinical manifestations (y-axis: neurologic, infectious, or autoimmune). (B) Percentage of patients who presented with various clinical features over the whole period of study until HSCT. MAS, macrophage activation syndrome; M.Still-like, Morbus Still-like.

All 46 patients (20 male, 26 female) had low (<5%) or no PNP enzyme activity in erythrocytes and/or genetic mutations compatible with the diagnosis of severe PNP deficiency. Four patients who were diagnosed at birth underwent ET.³⁰ A total of 23 patients (50%) had available PNP genotyping, which revealed compound heterozygous (n = 10) and homozygous (n = 13) variants with exon 2 and c.172C>T p.(Arg58∗) being the most common variant (n = 2 homozygous and n = 4 heterozygous),^15,42 followed by a homozygous intronic variant c.286-18G>A in 5 patients^15,43 (Table 2).

In patients with symptoms, primary immunodeficiency mainly manifested as recurrent bacterial respiratory tract infections (33/41; 80%) and viral infections (22/41; 54%), mainly by varicella zoster virus (VZV) (15/41; 37%), which caused encephalitis, myocarditis, pneumonitis, shingles (12/41; 29%), or postvaccination manifestations (3/41; 7%). Symptomatic infections with cytomegalovirus (CMV; 3/41; 7%), Epstein-Barr virus (EBV; 2/41; 5%), and herpes simplex virus (HSV; 1/41; 2.4%) were less common. Fungal infections at presentation consisted primarily of mucosal Candida-induced thrush (9/41; 22%).

Neurologic manifestations before HSCT were primarily developmental delay (33/41; 80%), followed by spastic paresis (n = 17; 41%), muscular hypotonia (n = 14; 34%), ataxia (n = 11; 27%), tremor/chorea (n = 2; 5%), and speech delay (n = 3; 7%). Other neurologic findings included polymicrogyria (patient 6), and sensorineural hearing loss (SNHL) (patient 15). Siblings of the latter patients had polymicrogyria or SNHL without evidence of a PNP deficiency.

The rate of patients with neurologic symptoms/developmental delay increased to 88% (36/41) overall at the time of HSCT, including 6 patients with central nervous system (CNS) infections and severe systemic viral infections caused by VZV and CMV (patients 4, 14, 17, 20, 27, and 28) and, in addition, 1 patient with EBV-lymphoid granulomatosis (patient 43)

Before HSCT, 19.5% (8/41) of patients developed AID, including Evans syndrome (autoimmune hemolytic anemia [AIHA]/immune thrombocytopenia [ITP]) in 7% (3/41), AIHA in 7% (3/41), AIN in 5% (2/41), and macrophage activation syndrome with polyarthritis in 2.5% (1/41).

One adolescent patient (pat. 32) developed an EBV-driven diffuse large B-cell lymphoma (Table 1; Figure 1).

Transplantation procedures, GF, and CTs

A total of 46 patients received a primary HSCT after conditioning with reduced-intensity conditioning (RIC; 15/46; 33%) and myeloablative conditioning (MAC; 29/46; 63%) or with no conditioning (2/46; 4%). Donors included human leukocyte antigen (HLA)–matched family (13/46; including 1 umbilical cord blood [UCB]), haploidentical family (8/46), matched unrelated (18/46; 15/46 HLA-10/10, 3/46 HLA-9/10) and unrelated UCB donors (7/46; 3/46 HLA 6/6, 3/46 5/6 and 1/46 4/6 match).

The graft sources included bone marrow (22/46; 48%), peripheral blood stem cells (16/46; 35%), and UCB (8/46; 17%). For GVHD and rejection prophylaxis, 67% (31/46) patients received in vivo T-cell depletion with antithymocyte globulin (ATG), including rabbit ATG (7/46; 15%), rabbit anti–T-lymphocyte globulin (4/46; 9%), horse ATG (1/46; 2%), and with alemtuzumab (19/46; 41%) or posttransplant cyclophosphamide (2/46; 4%). No serotherapy was given in 35% (16/46) of patients. In vitro graft manipulations, including T-cell receptor (TCR) α-β/CD19 depletion (2/46) and CD34 selection (1/46), were performed in 7% (3/46) of patients.

CTs included 5 hematopoietic SCBs (4/5, without T-cell depletion; 1/5 with CD34 selection), 3 mesenchymal stem cell infusions, and 4 DLIs (1/4 with CD45RA depletion), which were administered between 47 and 240 days after the first HSCT (Table 3).

Table 3.

HSCT characteristics and complications

ID	Age at HSCT (mo)	Donor	HLA	Graft	T-cell depletion∗	Conditioning†	Serotherapy	GVHD prophylaxis	Acute GVHD	Chronic GVHD	Post-HSCT CT	Outcome (mo/d after HSCT)	Complications >HSCT	AID >HSCT
1‡	35	MMFD	3/6	BM	Yes	Bu 16 Cy 200 TT 20 (MAC)	None	None	2	NA		Died (d+49)	ADV
2	26	MFD	6/6	BM	No	Bu 20 Flu 200 (MAC)	None	CSA	No	No		Alive (mo+156)
3	51	MFD	6/6	BM	No	Bu 16 Flu 120 (MAC)	None	CSA	3	Yes		Alive (mo+125)
4‡	35	MUD	10/10	BM	No	Bu 19.2 Flu 160 Cy 120 (MAC)	None	CSA/MTX	2	Yes		Died (mo+33)	Sepsis/meningitis (chronic extensive GVHD)
5	14	MUD	10/10	BM	No	Bu 19.2 Flu 160 Cy 120 (MAC)	None	CSA/MTX	No	No		Alive (mo+84)
6	10	MMFD	3/6	PBSC	Yes	Bu 16 Flu 160 TT 10 (MAC)	None	None	No	No		Alive (mo+70)
7	27	MFD	6/6	BM	No	None	None	CSA	No	No		Alive (mo+221)	Disseminated BCG Psoas abscess	AIHA
8	16	MFD	6/6	BM	No	Bu 20 Cy 200 (MAC)	None	CSA	No	No		Alive (mo+151)
9	14	MFD	6/6	BM	No	Bu 16 Flu 160 (MAC)	None	CSA/MTX	No	No		Alive (mo+114)
10	25	MFD	6/6	BM	No	Flu 150 Mel 140 (RIC)	ATG-G 20	CSA	1	Yes		Alive (mo+161)
11	6	MFD	6/6	BM	No	Flu 150 Mel 140 (RIC)	ATG-G 20	CSA	3	Yes	DLI (d+73)	Alive (mo+98)	RTI Klebsiella, pancytopenia
12	72	MUD	10/10	PBSC	No	Bu 12.8 Cy 120 (MAC)	ATG-G 40	CSA	No	No		Alive (mo+126)
13	20	MMFD	4/6	PBSC	Yes CD34+pos	Flu 150 Mel 140 (RIC)	ATG-T 20	None	1	No	2nd HSCT (mo+14)	Alive (mo+260)		AIHA§ ITP§
14	47	MUD	10/10	BM	No	Bu 16 Cy 200 (MAC)	Alemt. 1	CSA/MTX	1	No		Alive (mo+192)	VZV (zoster)
15	39	MFD	6/6	PBSC	No	Flu 150 Mel 140 (RIC)	Alemt. 1	CSA/MMF	3	No		Alive (mo+137)	Pneumatosis intestinalis
16	2	MUD	10/10	BM	No	Bu 16 Cy 200 (MAC)	Alemt. 0.6	CSA/MTX	2	No	SCB (mo+9)	Alive (mo+240)	VOD, pneumonitis, nephritis	AIHA§ Alopecia totalis§
17	39	MFD	10/10	PBSC	No	Flu 150 Mel 140 (RIC)	Alemt. 1	CSA/ MMF	3	No		Alive (mo+65)
18	25	MFD	6/6	BM	No	Bu 16 Flu 160 (MAC)	None	CSA/MTX/ rituximab	No	No		Alive (mo+87)	VOD, pneumonitis
19	22	MUD	10/10	PBSC	No	Flu 150 Mel 140 (RIC)	Alemt. 1	CSA	No	No	2nd HSCT (mo+41)	Alive (mo+62)
20	168	MUD	10/10	PBSC	No	Flu 150 Mel 140 (RIC)	ATG-T 10	CSA/ MMF	No	No		Alive (mo+28)
21	28	UCB	5/6	UCB	No	Treo 42 Flu 150 (RIC)	Alemt. 0.3	CSA/ MMF	No	Yes		Alive (mo+20)
22	25	MFD	10/10	BM	No	Bu 16 Cy 200 (MAC)	None	CSA/ MTX	2	Yes		Alive (mo+125)
23	42	MFD	6/6	BM	No	Treo 42 Flu 160 TT 8 (MAC)	None	CSA/MTX	No	No		Alive (mo+22)
24	12	MUD	10/10	BM	No	Treo 42 Flu 160 (RIC)	ATG-G 45	CSA/MTX	No	No		Alive (mo+94)
25‡	31	MUD	10/10	BM	No	Flu 150 Mel 140 (RIC)	Alemt. 0.6	CSA/MMF	4	NA	SCB	Died (mo+8)	Acute relapsing GVHD (gut)
26	7	MUD	9/10	PBSC	Yes CD34+pos	Treo 36 Flu 150 (RIC)	Alemt. 1	CSA/MMF	1	No		Alive (mo+72)
27	36	MFD	6/6	UCB	No	Bu 16 Flu 150 (MAC)	None	CSA	No	No		Alive (mo+156)	VZV (zoster)
28	40	UCB	6/6	UCB	No	Bu 16 Cy 200 (MAC)	None	CSA/Pred	No	No		Alive (mo+96)
29‡	22	UCB	10/10	UCB	No	Bu 16 Flu 140 Mel 70 (MAC)	ATG-T 10	CSA/Pred	No	NA		Died (d+20)	Neurologic decline
30	20	MUD	10/10	BM	No	Bu cAUC 80 Flu 150 (MAC)	ATG-T 10	CSA/MTX	No	No		Alive (mo+108)	RTI (ADV, PIV3, metapneumovirus)
31	18	MUD	10/10	PBSC	No	Mel 140 Flu 150 (RIC)	Alemt. 1	CSA/MMF	1	Yes		Alive (mo+24)
32	192	MMFD	3/6	PBSC	Yes TCRab/CD19	Treo 42 Flu 150 TT 10 (MAC)	Alemt. 0.3	None	2	No		Alive (mo+28)	GI (ADV, NV) RTI (Covid-19)
33	43	UCB	5/6	UCB	No	Bu 16 Cy 200 (MAC)	ATG-E 90	CSA/pred	No	Yes		Alive (mo+156)		AIHA§ ITP§ DM Graves disease
34	119	UCB	4/6	UCB	No	Flu 125 Mel 140 (RIC)	Alemt. 2.3	Tacro MTX	No	No		Alive (mo+96)
35	92	UCB	6/6	UCB	No	Flu 125 Mel 140 (RIC)	Alemt. 2.4	Tacro/MTX			2nd HSCT (mo+5)	Autologous recovery	VZV (zoster) RTI by EBV
36‡	52	UCB	5/6	UCB	No	Flu 150 Mel 140 TT 7 (MAC)	Alemt. 3.2	Tacro/MTX	3	NA		Died (mo+8.5)	ADV, CMV (acute relapsing GVHD)
37	2	MMFD	7/10	BM	No	Bu cAUC 55 Cy 20 Flu 150 (RIC)	Alemt. 0.4	PtCY d+3/+4 Tacro/MMF	No	No	2nd HSCT (mo+17)	Alive (mo+80)	EBV (rituximab) RTI (Aspergillus)	AIHA
38	4	MUD	9/10	BM	No	Bu cAUC 79 Flu 160 (MAC)	Alemt. 0.6	CSA/MMF	3	No		Alive (mo+53)	VOD
39	41	MUD	10/10	PBSC	No	Bu 12 Flu 250 (MAC)	Alemt. 0.6	CSA/MTX	No	No		Alive (mo+30)	EBV+PTLD (rituximab) GI (sapovirus) Neisseria bacteremia
40	26	MUD	9/10	PBSC	No	Bu 12 Flu 250 (MAC)	Alemt. 0.6	CSA/MTX	2	No		Alive (mo+22)	EBV (rituximab) RTI (rhinovirus/enterovirus/CV) GI (sapovirus)
41	4	MUD	10/10	PBSC	No	Bu12 Flu 250 (MAC)	Alemt. 0.6	CSA/MTX	2	No	2nd HSCT (mo+15) (+SCB)	Alive (mo+32)	EBV (rituximab) RTI (enterovirus/rhinovirus, ADV)
42	12	MMFD	7/10	PBSC	No	None	None	PtCY d+3/+4 Tacro/MMF	No	No	SCB (mo+3)	Alive (mo+45)
43	22	MMFD	6/10	PBSC	Yes TCRab/CD19	Treo 42 Flu 150 Thio 10 (MAC)	ATG-T 5	Tocilizumab abatacept rituximab	No	No	CD45RA deplet. DLI (d+47, +81, +94)	Alive (mo+63)	RTI (HHV6, RSV) GI (RV, HHV6, ADV) Sepsis (Candida) Disseminated BCG	Arthritis
44‡	37	MUD	10/10	BM	No	Bu cAUC 75 Flu 160 (MAC)	ATG-T 7.5	CSA/MMF	No	NA		Died (d+15)	MAS
45	23	MMFD	5/10	PBSC	Yes TCRab/CD19	Treo 42 Flu 150 TT 10 (MAC)	ATG-G 30	MMF, pred, ruxolitinib, vedolizumab	Yes, skin, 2	No		Alive (mo+12)	EBV infection GI NV Zoster (VZV) RTI (rhinovirus/enterovirus, BV)
46	3	MUD	10/10	BM	No	Bu cAUC 79 Flu 180 (MAC)	Alemt. 0.6	CSA/MMF	No	No		Alive (mo+85)

ADV, adenovirus; Alemt., alemtuzumab (mg/kg body weight); ATG-E, horse anti–thymocyte globulin (mg/kg body weight); ATG-G, rabbit anti–T-lymphocyte globulin (mg/kg body weight); ATG-T, rabbit antithymocyte globulin (mg/kg body weight); Bu, busulfan (mg/kg body weight); BV, bocavirus; cAUC, cumulative area under the curve (mg/L × h); CSA, cyclosporine A; CV, coronavirus; Cy, cyclophosphamide; DM, diabetes mellitus; Flu, fludarabine (mg/m² body surface area); GI, gastrointestinal infection; Mel, melphalan; MFD, matched family donor; MMF, mycophenolate mofetil; MMFD, mismatched family donor; MTX, methotrexate; NA, not applicable; NV, norovirus; PBSC, peripheral blood stem cells; PIV3, parainfluenza virus type 3; Pred, prednisolone; PtCY, post-HSCT cyclophosphamide; PTLD, posttransplant lymphoproliferative disease; RTI, respiratory tract infection; Tacro, tacrolimus; TCRab, T-cell receptor αβ; Treo, treosulfan (g/m² body surface area); TT, thiotepa (mg/kg body weight); VOD, veno-occlusive disease; UCB, umbilical cord blood.

^∗In vitro depletion.

^†Definitions of conditioning (supplemental Material).

^‡Deceased patient.

^§Resolved after rituximab treatment.

One patient developed primary GF and 4 patients developed secondary GF, both occurring after MAC and RIC conditioning. There was no difference in the OS among those who received RIC conditioning,³⁴ those who received no conditioning,⁴⁴ and those who received MAC conditioning, but higher rates of a second HSCT (24%; 4/17) and additional secondary CT, eg, DLI (n = 1) and SCB (n = 2) (41%; 7/17), were observed in the RIC/no conditioning group, whereas the CT rate was lower in the MAC group (1 DLI, second HSCT, and SCB; 10%; 3/29).

All patients with GF underwent a second HSCT with 2 of the patients receiving grafts from the primary donors (haploidentical, matched unrelated donor [MUD]) and 3 patients receiving grafts from secondary donors (2 MUD, 1 haploidentical). For reconditioning chemotherapy, RIC (n = 4) and MAC (n = 1) conditioning regimens were administered. No GF or acute or chronic GVHD was observed after the second HSCT, and all patients survived (Table 3; supplemental Table 3).

GHVD and other complications

No unusual toxicities were reported after conditioning for the primary or secondary HSCTs with the exception of 2 episodes of nonlethal hepatic veno-occlusive disease. Acute GVHD grade 2 to 4 was observed in 13 (30%) patients and grade 3 to 4 was observed in 7 (16%) patients. Among the surviving patients, chronic limited GVHD was observed in 7 (16%) patients, and none had extensive GVHD.

Among the patients who died, 3 patients had either chronic extensive (patient 4) or acute relapsing GVHD (patients 25 and 36); 2 of them died as a consequence of infections (adenovirus and sepsis/meningitis) (Table 3).

A total of 24% (11/46) of patients had relevant infections, including viral infections, eg, adenovirus, human herpesvirus 6 (HHV6), CMV, EBV, VZV, rotavirus, sapovirus, enterovirus/rhinovirus, respiratory syncytial virus (RSV), human metapneumovirus, and SARS-CoV-2. Bacterial infections included Clostridium difficile, Klebsiella pneumoniae, viridans group streptococci, and bacillus Calmette-Guérin (BCG), whereas the fungal infections were caused by Aspergillus sp. and Candida sp. In 4 cases, EBV reactivation required rituximab administration. All of these infections resolved (Table 3).

Secondary AIDs were observed in 15% of patients (7/46), including AIHA (3/46) and Evans syndrome (2/46), Graves disease, macrophage activation syndrome, arthritis, alopecia totalis, and diabetes mellitus. All episodes of secondary autoimmune cytopenia resolved after intensified immunosuppression, including rituximab (Table 3; supplemental Table 3).

Survival

After a median follow-up period of 7.9 years (1.0-22.3), the 3-year OS and EFS probabilities were 86% (95% CI, 77-97) and 75% (95% CI, 64-89), respectively. A total of 40 patients survived, whereas 6 patients died (13%). Five of these deaths occurred because of infections, whereas 1 patient succumbed to status epilepticus shortly after undergoing HSCT. Four patients passed away during the first year following HSCT, whereas 2 patients died >2 years after HSCT (Figure 2A; Table 3).

Figure 2.

OS and EFS curves. Kaplan-Meier survival curves illustrating the OS following HSCT. (A) OS and EFS after HSCT. (B) OS based on age at neurologic presentation (NP) (<11 months vs ≥11 months). (C) OS stratified by time from diagnosis to HSCT (<24 months vs ≥24 months). (D) EFS based on donor type, namely matched family donor (MFD), mismatched family donor (MMFD), MUD. (E) OS by age at HSCT (<28 months vs ≥28 months). (F) OS according to conditioning regimen, namely MAC, RIC, or none.

MSD/matched family donor transplants produced 100% disease-free survival (DFS). Unrelated donor and cord blood transplantation exhibited slightly lower success rates with 80.7% of patients experiencing DFS. Five of the 6 deaths occurred after MUD/mismatched unrelated donor (3/6) or UCB (2/6) transplantation. Haploidentical transplants demonstrated an 87.5% DFS rate, although the number of patients in this group was limited (8/46).

Patients diagnosed at birth and who underwent early HSCT (median age of 4 months) exhibited superior survival outcomes (100%) when compared with the rest of the cohort (84% CI, 74-97; P = .28; supplemental Figure 1R), suggesting that early diagnosis and HSCT can improve the overall outcomes, similar to what is seen in patients with SCID.⁴⁵

Patients who underwent HSCT within 24 months after initial presentation demonstrated superior OS (95% CI, 87-100 vs 70% CI, 51-97; P = .049). Conversely, neurologic symptoms that occurred before 11 months of age were associated with reduced OS (69% CI, 49-96 vs 94% CI, 83-100; P = .027). Furthermore, patients who underwent HSCT at a younger age (before 28 months of age) exhibited improved OS (96% CI, 89-100 vs 73% CI, 56-97; P = .036). However, a contrasting trend toward poorer outcomes was observed in children who presented solely with neurologic symptoms (75% CI, 57-100 vs 92% CI, 83-100; P = .076). Graft source, gender, alkylator, RIC, MAC, time of initial clinical presentation, and presence of AID before HSCT did not have a significant influence on OS (Figure 2; supplemental Figure 1A-V).

Donor chimerism and immune restoration

At the last FU, all surviving patients had been weaned from immunosuppression and exhibited adequate T-cell numbers and function, which provided protection against opportunistic infections and cured secondary autoimmune cytopenia. In total, 85% of patients (34/40) have discontinued immunoglobulin replacement therapy, whereas 15% (6/40) patients remained on immunoglobulin replacement therapy; 4 of these patients have undergone previous rituximab treatment.

DC evaluation in whole blood was conducted at the last FU in 37 of the 40 (92,5%) surviving patients, including 5 patients who successfully underwent a second HSCT (supplemental Figure 3; supplemental Table 3). The DC analysis revealed that 68% (25/37) exhibited levels of >95%, 18% (7/37) demonstrated levels between 50% and 95%, and 14% (5/37) had levels of <50% (with at least 12%). The DC in CD15⁺ cells (neutrophils) was analyzed in 10 patients, revealing that 20% (2/10) exhibited levels of >95%, 20% (2/10) had levels of between 50% and 95%, and 40% (6/10) had levels <50% (with at least 5%). The analysis of DC in CD3⁺ cells (T cells) in 19 patients revealed that 42% (8/19) exhibited levels of >95%, 37% (9/19) demonstrated levels of between 50% and 95%, and 10.5% (2/19) showed levels <50% (at least 40%). The DC in CD19⁺/CD20⁺ (B cells) was analyzed in 13 patients, revealing that 31% (4/13) exhibited levels of >95%, 15% (2/13) had levels of between 50% and 95%, and 54% (7/13) had levels of <50% (with at least 3%).

Metabolic analyses were available in 67.5% (27/40) of patients of whom 9 had normalized urinary purine metabolites and 18 had normal or greater than 10% PNP enzyme activity. Our data were insufficient to correlate the normalization of urine metabolites and/or PNP enzymatic activity with a particular transplant approach (Table 4; supplemental Figure 3).

Table 4.

Donor cell chimerism and immune and metabolic parameters at last FU

ID	FU (mo)	DC WBC (%)	DC CD15+ (%)	DC CD3+ (%)	DC CD19+ (%)	PNP enzyme activity∗/metabolites†	CD3+ (per μL)	CD4+/naïve CD4+45RA+ (per μL)	CD8+ (per μL)	PHA (SI)	CD19 (per μL)	IVIG (yes/no)
1‡	Died
2	156	100	ND	ND	ND	ND	1655	750	750	228	325	No
3	125	100	ND	ND	ND	ND	1050	470	545	461	235	No
4‡	Died
5	84	100	ND	ND	ND	ND	2920	1570	990	1016	1100	No
6	70	100	ND	ND	ND	ND	655	370	260	251	240	No
7	221	50	22	93	ND	Metabolites normal	1843	684	1064	Normal	8	Yes
8	151	100	ND	100	92	Metabolites normal	2410	1353	780	Normal	1271	No
9	114	95	ND	100	ND	Metabolites normal	ND	ND	ND	ND	ND	Yes
10	161	40	ND	83	ND	ND	13 300	1260	11 080	Normal	280	No
11	98	90	ND	ND	ND	PNP enzyme normal	1796	949/92	847	ND	338	No
12	160	100	ND	ND	ND	ND	1160	499	549	ND	420	No
13§	268	100	ND	ND	ND	PNP enzyme normal	670	370/70	230	ND	120	No
14	192	35	11	85	ND	ND	1260	610/250	570	111	210	No
15	137	90	79	100	87	PNP enzyme normal	1340	720/430	530	220	170	Yes
16§	252	99	ND	ND	ND	PNP enzyme normal	1950	1546	371	516	Normal	Yes
17	65	ND	5	100	20	ND	3690	258	2331	Normal	111	No
18	87	100	ND	ND	ND	PNP enzyme normal	3691	1678	1918	Normal	881	No
19	62	48	ND	44	ND	ND	868	468	369	ND	233	No
20	28	99	ND	ND	ND	PNP enzyme normal	1335	427	836	ND	267	No
21	20	96	ND	ND	ND	ND	ND	ND	ND	ND	ND	No
22	125	100	ND	ND	ND	PNP enzyme normal	ND	ND	ND	ND	ND	No
23	112	100	ND	ND	ND	PNP enzyme normal	1796	929/543	683	Normal	521	No
24	94	70	ND	80	60	ND	2950	1621/1149	993	ND	315	No
25‡	Died
26	72.0	70	73	75	50	PNP enzyme normal	1650	750	570	235	280	No
27	156.0	100	ND	ND	ND	PNP enzyme normal	3063	950/260	582	Normal	1623	No
28	96.0	100	ND	ND	ND	PNP enzyme normal	1648	914/548	642	ND	345	No
29‡	Died
30	108	100	ND	ND	ND	PNP enzyme normal	3455	1462/950	1123	Normal	670	No
31	24	100	ND	ND	ND	ND	700	40	250	ND	340	Yes
32	28	100	ND	ND	ND	ND	normal	Normal	Normal		Normal	No
33	156	100	ND	ND	ND	PNP enzyme normal	1626	779/53	666	Normal	239	No
34	96	85	10	85	10	ND	341	203/30	124	ND	104	No
35	98	100	ND	100	100	ND	1636	1018/43	468	ND	650	No
36‡	Died
37	80	100	ND	ND	ND	Metabolites normal	2220	1110/701	832	0.95	462	No
38	53	ND	ND	ND	ND	Metabolites normal	3072	2035/1608	768	0.9	653	No
39§	30	100	100	100	100	Metabolites normal	2180	930	640	Normal	500	No
40§	22	99	98	95	96	Metabolites normal	980	252	340	Normal	260	No
41§	32	ND	5	77	3	Metabolites normal	1430	201	760	Normal		Yes
42	45	12	ND	80	6	Metabolites normal	272	114/11	209	Normal	167	No
43	63	100	ND	100		ND	Normal	Normal	Normal	ND	Normal	Yes
44‡	Died
45	12	100	ND	100	100	ND	4580	ND	1860	34.7	1090	No
46	85	77	ND	79	45	PNP enzyme normal	1760	ND	750	1171	310	No

IVIG, intravenous immunoglobulin dependent; ND, not done; Normal, testing of lymphocyte subpopulations, PHA mitogen stimulation, urinary purine metabolites within the normal range; PHA SI, phytohemagglutinin stimulation of lymphocytes; SI, stimulation index; WBC, white blood cells.

^∗Erythrocyte PNP enzyme activity in untransfused patients.

^†Urinary purine metabolites (uric acid, deoxyguanosine triphosphate, guanosine, inosine, deoxyguanosine, and deoxyinosine).

^‡Deceased patient.

^§Treated with rituximab.

Neurologic outcomes

At the most recent FU, the treating physician evaluated the severity of neurologic symptoms and the CHIMO score and classified the results into 3 categories, namely improved (17/40; 42.5%), clinically stable (n = 16/40; 40%), or declined (n = 7/40; 17.5%). The median CHIMO score was determined to be 14 (range, 6-17), with medians of 3 (0-3) for cognition, 3 (1-3) for hearing, 4 (1-4) for interaction, 3 (1-4) for movement, and 2 (0-3) for occupation. The median Karnofsky and Lansky scores were 90% and 85%, respectively, with a range of 40% to 100% for both scores (Table 5; Figures 3 and 4). In most of our patients, no further neurologic deterioration occurred after HSCT once adequate engraftment of donor myeloid cells had been achieved. After a median FU period of >7 years following HSCT, most surviving patients had reached preschool/school age, thereby enabling more precise categorization of the category occupation within the CHIMO score. The analysis of CHIMO scores revealed that 50% (20/40) of patients attained the highest scores of between 15 and 17, whereas an additional 35% (14/40) scored within the range of 12 to 14. Conversely, 15% (6/40) of patients, were in the lowest score range of 6 to 11. All 6 patients who were diagnosed early by family history and who underwent early HSCT were alive and were assessed with a median CHIMO score of 14 (9-17) at a median FU of 63 months (Table 5), which was comparable with the median CHIMO score of 14 for the entire cohort. Four of these early-diagnosed patients received ET shortly after birth, which was reported to achieve measurable purine metabolite detoxification before HSCT.³⁰ Although early detoxification until HSCT seems a rational approach for early-diagnosed patients with PNP,²⁷ it was as yet impossible to determine whether this affects the long-term neurologic outcomes given the limited number of subjects.

Table 5.

Neurologic outcome and CHIMO score at last FU

ID	FU (mo)	Cognition (score)	Hearing (score)	Interaction (score)	Movement (score)	Occupation (score)	CHIMO (score)	Karnofsky (score)	Lansky (score)	Neurologic evaluation
1∗	Died
2	156	3	3	4	3	3	16	100	100	Stable
3	125	2	3	3	4	3	15	100	100	Stable
4∗	Died
5	84	0	3	1	1	1	6	40	40	Stable
6	70	0	3	2	1	0	6	50	50	Stable
7†	221	3	3	4	2	2	14	50	70	Improved
8	151	3	3	4	4	3	17	100	100	Improved
9	114	3	3	4	3	3	16	90	100	Stable
10	161	1	2	3	1	1	8	50		Stable
11	98	2	3	3	3	2	13		100	Improved
12	160	3	3	3	4	3	16	100		Stable
13	268	3	3	4	3	3	16	90	90	Stable
14	192	3	3	4	1	2	13	80	90	Improved
15	137	3	1	4	4	3	15	100	100	Stable
16†	252	2	3	3	1	2	11	40	100	Declined
17	65	2	3	3	2	2	12		70	Stable
18	87	3	3	4	4	3	17	100	100	Improved
19	62	1	3	3	3	2	12	70	100	Improved
20	28	3	3	4	3	2	15	90		Improved
21	20	2	3	3	3	2	13	70		Improved
22	125	3	3	4	4	3	17	100		Stable
23	112	3	3	3	3	3	15		80	Improved
24	94	3	3	4	3	3	16		90	Declined
25∗	Died
26†	72.0	1	2	3	1	2	9	50	75	Stable
27	156.0	3	3	4	3	3	16	90		Improved
28	96.0	1	3	4	2	2	12	100		Declined
29∗	Died
30	108	1	3	4	2	2	12	70		Stable
31	24	2	3	4	3	3	15	90	90	Improved
32	28	2	3	3	2	2	12	60		Stable
33	156	1	3	4	3	2	13	90		Stable
34	96	3	3	4	4	3	17	100		Improved
35	98	3	3	4	2	3	15	90		Improved
36∗	Died
37†	80	3	3	3	4	3	16		100	Declined
38†	53	3	3	4	4	3	17		100	Stable
39	30	3	3	4	4	3	17		100	Improved
40	22	3	2	3	4	3	15		100	Improved
41†	32	3	3	3	2	3	14		90	Declined
42	45	1	3	1	3	1	9	70	90	Declined
43	63	3	3	4	1	2	13	60	70	Improved
44∗	Died
45	12	2	3	3	3	2	13	50	70	Improved
46†	85	3	3	3	2	2	13	40	70	Declined

FU, follow-up (in months after HSCT).

^∗Deceased patient.

^†Perinatal/antenatal diagnosis owing to family history (patients 37, 38, 41, and 46 had received red cell exchange transfusions after birth).

Figure 3.

Distribution of CHIMO scores at last FU visit. Plot illustrating the distribution of CHIMO scores across different domains, namely cognition, hearing, interaction, movement, and occupation, as well as the overall CHIMO score. Scores are displayed for individual patients at their specific age at their respective time of FU for the assessment of the CHIMO score.

Figure 4.

Severity of categorized symptoms before and after HSCT. Heat map depicting the severity scores of clinical symptoms before and after HSCT divided into the categories, namely neurology, infection, and autoimmunity. Scores were based on clinical descriptions and CHIMO scores (refer to the supplemental Material). Asterisk (∗) marks the 6 patients diagnosed perinatally by family history.

Discussion

The initial cases of PNP deficiency were reported in 1975, and since then, ∼100 patients with PNP deficiency have been described in the literature,^1,7,8,15 primarily in the form of case reports. In 1991, only a minority (29%) survived.¹ Between 1975 and 2022, only 22 patients have been reported to have undergone HSCT.¹⁵ Long-term studies on the clinical and neurologic outcomes are still lacking, and so assessing the long-term prognosis of these patients after HSCT remains a major challenge today.^{3,4,11,15,20,23,24,28,29,46, 47, 48, 49, 50}

Early diagnosis of PNP deficiency in infancy is challenging because of the variability in symptoms of immunodeficiency, which are more consistent with CID than SCID. For instance, no cases of Pneumocystis jirovecii pneumonia have been documented in our or other series.^15,41,45 The most prevalent infectious symptoms during infancy were nonspecific respiratory tract infections and thrush, whereas severe or atypical VZV infections, including from vaccine strains, were observed exclusively in patients beyond infancy. Furthermore, approximately one-third of our patients exhibited neurologic abnormalities without any clinical signs of immunodeficiency (see also Habib Dzulkarnain et al¹⁵; Torun et al¹⁷; Tsui et al²⁷; Dror et al^{15, 17, 27, 51}). Motor delay manifested as global hypotonia in infants and as spastic paresis and ataxia in older children, whereas secondary AIDs were mainly observed late in infancy or in the second year of life. Consequentially, the median age at HSCT in our cohort was remarkably late at 26 months of age. Before HSCT, the percentage of patients with neurologic abnormalities had more than doubled by the time of HSCT, often because of infectious complications that involved the CNS. This underscores the importance of thorough clinical evaluation for timely diagnosis.

Magnetic resonance imaging of the CNS, which was not standardized in this study, showed many normal but also occasional pathologic findings, including atrophy of the corpus callosum and delayed myelination. Similar inconsistent findings have been documented in previous studies of patients with PNP deficiency.^{4,13,15,17,52} However, in PNP-deficient mice, the small size of the cerebellum, corpus callosum, and thalamus has been attributed largely to increased neuronal apoptosis, which does not seem to be as clearly demonstrable in humans.⁵³ Conversely, other clinical neurologic abnormalities, including microcephaly/brachycephaly and SNHL were clearly not caused by PNP deficiency, because these same findings were observed in consanguineous families in siblings without a detected PNP mutation. In contrast with the manifestations associated with adenosine deaminase deficiency, no additional cases of SNHL were observed,¹⁵ suggesting that this complication is unlikely in PNP deficiency.

Conditioning regimens A/B/C of the ESID/EBMT guidelines comprising MAC and reduced-toxicity regimens³⁴ were overall successful in the present PNP-deficient cohort. No excessive endothelial, skin/epithelial, or organ toxicity was observed after MAC conditioning, suggesting that the increased radiosensitivity of thymocytes and peripheral T cells demonstrated in PNP-deficient mouse models is clinically irrelevant in affected humans.^2,51 The necessity of brain conditioning with busulfan to facilitate the migration of monocytic precursors into the CNS and their possible subsequent transdifferentiation into neuroglial cells is a subject of debate in PNP deficiency. Experiments have demonstrated that the proportion of glial precursors from PNP-deficient induced pluripotent stem cells is not statistically different from that observed in healthy controls. This observation suggests that glial cells may not play a significant role in the neurologic impairment associated with PNP deficiency.²⁷ The present study supports this finding by demonstrating no discernible differences in neurologic outcomes between patients who received busulfan-containing regimens and those who received treosulfan, which does not penetrate the blood-brain barrier well. Because of a higher rate of CTs after RIC regimens, which were mainly based on melphalan/fludarabine (regimen E³⁴), we believe that regimens A/B/C³⁴ are preferable in patients with PNP deficiency to ensure long-term myeloid donor engraftment,⁴⁴ but larger studies are needed to prove this.

Despite the use of anti–T-cell–mediated serotherapy in most patients, complications such as chronic GVHD, GF, and secondary AIDs+ were not uncommon. In order to further optimize conditioning regimens and reduce these complications, it is essential to continue refining therapeutic drug monitoring of busulfan,⁵⁴ treosulfan,⁵⁵ ATG,⁵⁶ or alemtuzumab,⁵⁷ as well as T-cell depletion techniques, such as in vitro TCR α-β/CD19-depletion⁵⁸ or cyclophosphamide after haploidentical HSCT.

T-cell lymphopenia resolved in all surviving patients in our cohort, although a few patients had stable but low levels of mixed myeloid DC. No new neurologic symptoms developed in these patients with low-level DC, suggesting that residual PNP activity, even if generated by a small number of engrafted donor myeloid and erythroid progenitor cells, may lead to immunologic and metabolic recovery. Despite the limitation that biochemical analyses were not consistently available for all patients in the present report, patients with mixed myeloid DC were shown to have sufficient PNP enzyme activity and/or the absence of urinary purine metabolites. Our findings are consistent with reports of patients with partial PNP deficiency and residual PNP enzyme activity (8%-11%) who had near-normal immunity and normal neurologic development into their third decade of life.² Furthermore, measurable detoxification has already been demonstrated in a patient with 5% myeloid DC after unconditioned HSCT.⁴⁴

Assessment of the neurologic outcomes after HSCT remains challenging, particularly in infants and young children who reach new developmental milestones over time.^26,48,59 The spectrum and variable degree of neurologic impairment in individuals with PNP deficiency could hypothetically be caused by the interindividual variability in residual enzyme activity. However, we were unable to confirm this hypothesis in our cohort, because we did not observe any differences in the outcomes between patients with absent or residual enzyme function at baseline. The presence of reported differences in neurologic phenotypes, even among family members with identical mutations, suggests that additional factors may play an important role.²⁷ These additional factors could include interindividual differences in the susceptibility to neuronal apoptosis and external factors, such as secondary infections of the CNS.

To more precisely assess neurologic outcomes, we included the CHIMO score in the outcome analysis with the caveat that although all CHIMO subscores had been validated, the overall CHIMO score had not.^{36, 37, 38} The median CHIMO score of 14 achieved by the entire surviving cohort was compatible with satisfactory academic and professional performance, as well as the ability to live independently without external assistance in adulthood. Residual motor impairment was more common than severe cognitive impairment with the former very rarely requiring the use of a wheelchair. A detailed analysis of observed behavioral problems, such as attention deficit or autism spectrum disorders, which are quite prevalent in adenosine deaminase deficiency, was beyond the scope of this analysis. However, these conditions may have contributed to educational challenges and lower vocational performance in some patients.

This investigation of the largest cohort of patients with PNP to date confirms that HSCT, using blood stem cells from HLA-matched and haploidentical donors, is curative for affected patients after using different conditioning regimens. Survival with sufficient myeloid DC was associated with metabolic correction and stabilization of neurodevelopmental status without further neurologic decline, which led to predominantly satisfactory long-term neurologic outcomes.

Although timely diagnosis and HSCT improved the OS, we were unable to assess the impact of disease-specific factors on neurologic outcomes, such as individual susceptibility to neuronal apoptosis,²⁷ or additional damage caused by CNS infections¹⁸ because of the limitations of our study. In addition, we were unable to show that patients who were diagnosed at birth and were neurologically asymptomatic³⁰ achieved superior median CHIMO scores when compared with patients who were diagnosed later upon clinical symptoms. Given the limited number of neurologically asymptomatic patients, the need for larger comparative studies with early-diagnosed patients with PNP deficiency is mandatory.

The introduction of a sensitive newborn screening method for PNP deficiency could therefore be useful to commence detoxification and HSCT on affected patients as early as possible. Future studies will show whether this can further improve the results presented here.

Conflict-of-interest disclosure: I.M. reports being part of a European Reference Network for Rare Immunological and Autoimmune Diseases Core Center; is a senior clinical researcher at Research Foundation–Flanders; and is supported by the Jeffrey Modell Foundation. T.G., M.F., M.H.-H., U.Z., and A.H. report being part of the Children’s Research Center of the University Children’s Hospital, Zürich, Switzerland. The remaining authors have no conflict of interest to disclose.

A complete list of the members of the EBMT Inborn Errors Working Party appears in the supplemental Appendix.