REVIEW

A Review of Chronic Granulomatous Disease

Danielle E. Arnold . Jennifer R. Heimall

Received: September 15, 2017 / Published online: November 22, 2017� The Author(s) 2017. This article is an open access publication

ABSTRACT

Chronic granulomatous disease (CGD) is a pri-mary immunodeficiency caused by defects inany of the five subunits of the NADPH oxidasecomplex responsible for the respiratory burst inphagocytic leukocytes. Patients with CGD are atincreased risk of life-threatening infections withcatalase-positive bacteria and fungi and inflam-matory complications such as CGD colitis. Theimplementation of routine antimicrobial pro-phylaxis and the advent of azole antifungals hasconsiderably improved overall survival. Never-theless, life expectancy remains decreased com-pared to the general population. Inflammatorycomplications are a significant contributor tomorbidity in CGD, and they are often refractoryto standard therapies. At present, hematopoieticstem cell transplantation (HCT) is the onlycurative treatment, and transplantation out-comes have improved over the last few decadeswith overall survival rates now[90% in childrenless than 14 years of age. However, there remainsdebate as to the optimal conditioning regimen,

and there is question as to how to manage ado-lescent and adult patients. The current evidencesuggests that myeloablative conditioning resultsis more durable myeloid engraftment but withincreased toxicity and high rates of graft-ver-sus-host disease. In recent years, gene therapyhas been proposed as an alternative to HCT forpatients without an HLA-matched donor. How-ever, results to date have not been encouraging.with negligible long-term engraftment ofgene-corrected hematopoietic stem cells andreports of myelodysplastic syndrome due toinsertional mutagenesis. Multicenter trials arecurrently underway in the United States andEurope using a SIN-lentiviral vector under thecontrol of a myeloid-specific promoter, and,should the trials be successful, gene therapymaybe a viable option for patients with CGD in thefuture.

Keywords: Chronic granulomatous disease;Gene therapy; Hematopoietic stem celltransplantation; Infections; Inflammation;Treatment

INTRODUCTION

Chronic granulomatous disease (CGD) is aninherited primary immunodeficiency caused byfunctional impairment of the NADPH oxidasecomplex in neutrophilic granulocytes and

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D. E. Arnold � J. R. Heimall (&)Children’s Hospital of Philadelphia, Wood Center,Rm 3301, 3401 Civic Center Blvd, Philadelphia, PA19104, USAe-mail: heimallj@email.chop.edu

Adv Ther (2017) 34:2543–2557

DOI 10.1007/s12325-017-0636-2

monocytes and characterized by recurrent andsevere infections, dysregulated inflammation,and autoimmunity. The NADPH oxidase com-plex is comprised of both membrane-bound andcytosolic proteins that function in concert uponphagocyte activation to produce reactive oxy-gen species (ROS) essential for the normal kill-ing of bacteria and fungi [1]. The catalyticglycoprotein gp91phox and non-glycosylatedprotein p22phox are located in the cell mem-brane and together form the heterodimer cyto-chrome b558. Upon phagocyte activation, thecytosolic proteins p47phox, p67phox, and p40phox

translocate to cytochrome b558 and recruitRac1/2. This results in a conformational changein gp91phox, which enables cytosolic NADPH todonate an electron to molecular oxygen in thephagolysosome to form superoxide ions.Superoxide ions are then used to generate ROSsuch as hydrogen peroxide, hypochlorous acid,hydroxyl radicals, and secondary amines thatare highly toxic to phagocytosedmicroorganisms.

Mutations in any of the five structural sub-units of the NADPH oxidase complex result indefective ROS production and the syndrome ofCGD. The transmembrane glycoproteingp91phox is encoded by CYBB on the X chro-mosome and accounts for approximately two--thirds of cases of CGD. Autosomal recessivemutations in NCF1 (p47phox) account for about20% of cases, and mutations in CYBA (p22phox)and NCF2 (p67phox) each account for about 5%of cases [2–6]. There has been one reported caseof an NCF4 (p40phox) mutation resulting inCGD [7]. The incidence of CGD in the UnitedStates and Europe is around 1 in 200,000 to 1 in250,000 live births [2, 5]. However, the inci-dence varies significantly worldwide, from 1:1million in Italy to 1:70,000 in the Israeli Arabpopulation [4, 8], and, in countries with highrates of consanguinity, the rate of autosomalrecessive CGD exceeds that of X-linked CGD[5, 8–10].

CGD may present at any age from infancy tolate adulthood; however, the vast majority ofpatients are diagnosed at less than 5 years of age[3–5]. In general, patients with X-linked CGDhave a more severe disease course with earlierage at presentation and earlier age of death

[2, 5]. Mechanistically, the survival of patientswith CGD is strongly associated with residualsuperoxide production independent of thespecific gene affected [6].

CGD was initially described as ‘‘a fatal gran-ulomatous disease of childhood,’’ and. histori-cally, most patients with CGD died by 10 yearsof age [11]. However, improved awareness of thedisease and advances in management have ledto a marked improvement in life expectancy.Before the introduction of oral antifungals,Winkelstein et al. reported a mortality rate of5% per year for X-linked CGD and 2% per yearfor autosomal recessive CGD [2]. More recentstudies now report a survival rate of approxi-mately 90% at 10 years of age, which has beenattributed to improved recognition and earlydiagnosis leading to earlier therapies, includingmore efficacious antimicrobial prophylaxis, useof interferon-gamma (IFN-c) supplementationfor infection prophylaxis, and use ofhematopoietic stem cell transplantation (HCT)[6]. Nevertheless, the median age of deathremains around 30–40 years, and patients tendto become increasingly debilitated with poorquality of life with advancing age [3–5, 12].

This review aims to summarize the clinicalphenotype of CGD, including infectious andinflammatory manifestations, and to update thecurrent data on conventional management,HCT, and gene therapy. It also aims to identifyquestions that remain with regard to optimalmanagement of patients with CGD, particularlywith respect to HCT. This article is based onpreviously conducted studies and does notinvolve any new studies of human or animalsubjects performed by any of the authors.

INFECTIONS

Infections are primarily with a subset of cata-lase-positive microorganisms, and the mostcommon sites of infection are the lungs, skin,lymph nodes, and liver. In North America andEurope, the most frequent pathogens areAspergillus spp., Staphylococcus aureus,Burkholderia cepacia, Serratia marcescens, Nocar-dia spp., and Salmonella [1–5, 12]. In developingcountries, Bacille Calmette-Guerin (BCG) and

2544 Adv Ther (2017) 34:2543–2557

Mycobacterium tuberculosis are important patho-gens [8, 13, 14]. There are a number of unusualbacteria that have been reported over the lastfew decades that are virtually pathognomonicfor CGD. Chromcobacterium violaceum and Fran-cisella philomiragia are found in brackish waterand most frequently cause skin and deep tissueabscesses and sepsis in CGD [15–17]. Gran-ulibacter bethesdensis is a Gram-negative rod thatcauses chronic necrotizing lymphadenitis andsepsis [18], and Burkholderia gladioli has beenreported as a cause of osteomyelitis and sepsis[19, 20]. Infection with any of these microor-ganisms should prompt evaluation for CGD.

CGD has the highest prevalence of invasivefungal infections among all primary immun-odeficiencies, affecting 20–40% of CGDpatients, and invasive fungal infections remainan important contributor to morbidity andmortality. [12, 21–23]. The lungs and chest wallare the most common sites of infection, andAspergillus fumigatus followed by A. nidulans arethe most commonly isolated pathogens[12, 21–23]. A. fumigatus was previously theleading cause of mortality in CGD; however,with the advent of azole antifungal treatment,death from A. fumigatus is now uncommon [23].Conversely, A. nidulans infections cause moresevere, refractory, and invasive disease withhigh mortality rates [21–23]. Notably, the inci-dence of A. nidulans infections has increasedsince widespread implementation of itracona-zole prophylaxis. Other Aspergillus spp, includ-ing A. viridinutans, A. tanneri, and Neosartoryaudagawae also cause disease in CGD and aredifficult to treat [24–26] After Aspergillus spp.,Rhizopus spp. and Trichosporon spp. are the mostcommonly identified fungal pathogens in CGD[27]. Other rare fungi seen in patients with CGDinclude Paecilomyces variotii, Paecilomyces lilaci-nus, Phellinus tropicalis, and Geosmithia argillacea[28–32]. Mulch pneumonitis deserves specialmention, as it is exclusive to CGD and is asso-ciated with a high rate of mortality if notidentified early. Mulch pneumonitis is due to anexuberant inflammatory response to fungalelements in aerosolized decayed organic matterand should be considered in all cases of unex-plained pneumonitis in previously well patients[33, 34]. Of note, dimorphic mold infections

such as histoplasmosis and blastomycosis andthe yeast infection cryptococcosis are not seenin CGD. Mucormycosis is also rare in CGD andonly occurs in the setting of significantimmunosuppression [35].

INFLAMMATORY COMPLICATIONS

In addition to recurrent and severe infections,dysregulated inflammation is commonly seenin CGD patients. A recent study on a Frenchcohort of 98 patients reported inflammatorymanifestations in 69.4% of patients, and themost commonly affected organs were the GItract (88.2% of patients), lungs (26.4%), uro-genital tract (17.6%), and eyes (8.85%) [36].About 10% of patients also had autoimmunecomplications. Patients with X-linked CGD hadtwo times the rate of inflammatory complica-tions compared to patients with autosomalrecessive CGD.

GI tract manifestations are common, with areported incidence ranging from 33% to 60% ofpatients with CGD [36, 37]. Symptom onsetmay be at any time, but most affected patientsdevelop GI involvement in the first decade oflife [37]. Importantly, GI manifestations mayprecede the diagnosis of CGD and the devel-opment of infectious complications. As such,CGD should be considered in all patients whopresent with early onset inflammatory boweldisease. GI symptoms are generally non-specificand include abdominal pain, noninfectiousdiarrhea, oral aphthae, nausea and vomiting,and failure to thrive [36–38]. The colon is themost frequently affected site, and patients withCGD are particularly prone to developing peri-anal disease with high rates of anal fistulae andperirectal abscesses [37–40].

In addition to inflammatory bowel disease,liver involvement is frequent and can be sig-nificant. Patients with CGD may developnodular regenerative hyperplasia, non-cirrhoticportal hypertension, hepatosplenomegaly, andsplenic sequestration [41]. Liver manifestationsare often progressive, and, notably, the devel-opment of thrombocytopenia secondary tosplenic sequestration is a strong predictor ofmortality [42]. Genitourinary tract

Adv Ther (2017) 34:2543–2557 2545

manifestations are common and include blad-der granulomata, ureteral obstruction, and uri-nary tract infections, especially in patients withgp91phox and p22phox deficiency [43]. Eosino-philic cystitis has also been reported in childrenwith CGD [44, 45]. Pulmonary manifestationsmay include granulomatous lung disease andinterstitial pulmonary fibrosis [36, 38]. Ocularinvolvement with chorioretinitis, uveitis, andocular granulomata has been reported [36]. Ofnote, macrophage activation syndrome has alsobeen reported in CGD patients and is a poten-tially life-threatening inflammatory complica-tion [46, 47].

X-LINKED CARRIERS

Female carriers of X-linked CGD have a dualphagocyte population due to lyonization, andcases of severe skewing of X-chromosomeinactivation have been reported in whichpatients are at risk for CGD-type infections.Reports of female carriers with discoid lupuserythematosus, photosensitivity rashes, andother autoimmune phenomena have beenpublished [48, 49]. Inflammatory bowel diseasehas also been reported in women with skewedX-inactivation [50].

A recent UK survey of 94 female carriers ofX-linked CGD demonstrated that these indi-viduals may be more symptomatic than previ-ously thought [51]. Cutaneous symptoms werereported by 63 (79%) women, most frequentlyphotosensitivity but also malar-like lupus rashand eczema; skin abscesses were reported by 14(17%) women; gastrointestinal symptoms werereported by 40 (42%) women; and 24 (26%)women met criteria for systemic lupus erythe-matosus. Female carriers with skin abscesses andchronic diarrhea were found to have a signifi-cantly lower neutrophil respiratory oxidativeburst than unaffected carriers. Interestingly,there seemed to be no relationship betweenautoimmunity and neutrophil respiratoryoxidative burst.

Another recent study from the NIH of 162female carriers of X-linked CGD again showedgreater symptomatology than previously rec-ognized, although not to the same extent as the

UK survey [52]. In the NIH study, 25% ofwomen had cutaneous symptoms, and 19% hadautoimmunity. Fourteen women (15%) had ahistory of severe infection caused by typicalCGD pathogens. There was a clear correlationbetween history of severe infection and percentof neutrophils with normal oxidative capacity.Women with less that 20% normal neutrophiloxidative capacity had increased infections, andless than 10% was highly associated with severeinfection. As in the UK survey, there was noassociation between neutrophil oxidativecapacity and symptomatic autoimmunity.

These findings create questions regarding thelong-term health of female carriers and theirsuitability for consideration as donors for HCTin their affected family members, since thedegree of lyonization can change over time.

CONVENTIONAL MANAGEMENT

Conventional management is predominantlywith lifelong antibiotic and antifungal prophy-laxis. Trimethoprim-sulfamethoxazole has beenshown to reduce the incidence of bacterialinfections from 15.8 to 6.9 infections per 100patient-months in patients with X-linked CGDand from 7.1 to 2.4 per 100 patient-months inpatients with autosomal recessive CGD [53].Itraconazole prophylaxis was shown to be welltolerated in a trial of 39 patients randomized toreceive either placebo or itraconazole. Only onepatient receiving itraconazole had a seriousfungal infection compared to seven in the pla-cebo group [54]. As such, lifelong prophylaxiswith trimethroprim-sulfamethoxazole (5 mg/kg/d div BID up to 320 mg trimethoprim a day)and itraconazole (5 mg/kg/d up to 200 mgdaily) is recommended. Dicloxacillin andciprofloxacin are options for patients with sul-famethoxazole allergy or G6PD deficiency. Forthose unable to tolerate itraconazole,posaconazole has been shown to be safe andeffective [55]. Patients with CGD should receiveall routine childhood immunizations except forthe BCG vaccine.

The prophylactic use of IFN-c remains vari-able. A large randomized, double-blind,placebo-controlled study of 128 patients with

2546 Adv Ther (2017) 34:2543–2557

CGD showed a clear benefit of IFN-c prophy-laxis with a decrease in both the number andseverity of infections (14/63 patients assigned toIFN-c developed serious infections during thestudy period versus 30/65 patients assigned toplacebo) regardless of inheritance pattern, sex,or use of antibiotic prophylaxis [56]. Long-termfollow-up of 9 years demonstrated sustainedbenefit [57]. Conversely, a prospective Italianstudy showed that long-term prophylaxis withIFN-c did not significantly change the rate oftotal infection per patient-year compared tocontrol, and the group determined there was noevidence to justify long-term prophylaxis withIFN-c [4]. At our institution, the decision forIFN-c prophylaxis is made on a case-by-casebasis, and is particularly encouraged for patientsexperiencing increased infections.

Several large studies have reported rates ofinfection of around 0.3 per year despite appro-priate antimicrobial prophylaxis [4, 12]. Infec-tions should be treated early and aggressively,and initial antibiotic therapy should providestrong coverage for both S. aureus andGram-negative bacteria, including B. cepacia(e.g., combination of vancomycin/clin-damycin/oxacillin and ceftazidime/carbapenemdepending on local resistance patterns). Treat-ment strength dosing of trimethoprim-sul-famethoxazole to cover ceftazidime-resistant B.cepacia and Nocardia should also be consideredas part of the initial empiric therapy. If patientsdo not improve within 24–48 h, then moreaggressive diagnostic procedures to identify theresponsible pathogen should be considered.

There should always be a high index of sus-picion for invasive fungal infection in patientswith CGD. Invasive fungal infections mostcommonly affect the lungs and chest wall. Inpatients with pulmonary symptoms and/orfever of uncertain origin, antifungal therapyshould be initiated as part of the initial empirictherapy. Of note, Aspergillus serological tests(e.g., Aspergillus galactomannan) and bronchialalveolar lavage have particularly low sensitivityin patients with CGD and should not be reliedupon for diagnosis [23]. Similarly, the efficacy of1,3-b-D-glucan testing is unclear in CGD. Vori-conazole is recommend as first-line therapy forits activity against Aspergillus spp. For infections

refractory to voriconazole, liposomal ampho-tericin B, caspofungin, posaconazole, or somecombination thereof may be considered. Surgi-cal intervention is often necessary, and patientsgenerally require prolonged treatment courses.Rescue HCT and/or gene therapy have beenproposed as viable options for life-threateninginfections resistant to antifungal treatment.

Corticosteroids have traditionally beenavoided in patients with CGD and activeinfection; however, a number of reports indi-cate that steroids may be used in conjunctionwith appropriate antimicrobials to treat hyper-active inflammation. Liver abscesses affectabout one-third of patients with CGD, and theyare often recurrent [12, 58]. Liver abscesses aredense, caseous, and difficult to drain and fre-quently require surgical intervention. However,in a case series of nine patients at the NIH withStaphylococcal liver abscesses refractory to con-ventional therapy, the addition of corticos-teroids led to the successful resolution of liverabscesses without need for surgical intervention[59]. Corticosteroids have also been shown toplay a role in the treatment of respiratoryinfections, including Nocardia pneumonia andmulch pneumonitis [33, 60, 61].

Treatment of CGD colitis is often long-termand difficult. Patients typically respond to ster-oids, but relapse is common [37]. Treatmentwithinfliximab also leads to rapid improvement;however, it is associated with increased infec-tions and death in patients with CGD [62]. Assuch, TNF-a inhibitors should be strictly avoi-ded. Steroid-sparing agents used with varyingdegrees of success include salicylic acid deriva-tives, antimetabolites such as azathioprine, and6-mercaptopurine. IL-1R blockade using Ana-kinra resulted in rapid and sustained improve-ment of colitis in two patients with CGD [63]. Ofnote, hematopoietic stem cell transplantation iscurative, and most patients have complete reso-lution of colitis following transplantation.

HEMATOPOIETIC STEM CELLTRANSPLANTATION

Allogeneic HCT is the only curative treatmentfor CGD and may reverse both infectious and

Adv Ther (2017) 34:2543–2557 2547

inflammatory complications. However, patientswith CGD are prone to graft failure, and priorinfections and organ dysfunction may increasetransplant-related complications. Early reportsshowed that HCT was possible, but outcomeswere poorer than reported for other primaryimmunodeficiencies with a high rate of mor-tality, graft failure, and low donor chimerism[64]. These complications were at least partlyattributed to the use of reduced intensity con-ditioning (RIC), and it was proposed that moremyeloablative conditioning (MAC) may benecessary for stable engraftment. However,higher intensity conditioning typically resultsin more prolonged immunosuppression leadingto an increased risk of infection and require-ment for blood/platelet transfusions. HCT isalso associated with risk of developinggraft-versus-host disease (GVHD).

In 2002, Seger et al. published results from acohort of 27 European patients ranging from 3to 38.7 years of age who underwent HCT forCGD between 1985 and 2000 [65]. Twenty-fiveof the 27 patients received grafts from anHLA-identical sibling, and all but four patientsreceived MAC with busulfan and cyclophos-phamide. The four patients who received RICwere severely debilitated and not candidates forMAC. Overall survival was reported at 85% withan event-free survival of 81%. Notably, allpatients without severe active infection orinflammation did extremely well (18/18patients survived). Conversely, all four patientswith active fungal infections died, and thosewith active inflammation had high rates ofGVHD. Also of note, only two of the fourpatients who received RIC had stable myeloidengraftment. Ultimately, this study showed thatHCT with MAC was a viable option for patientswith CGD and an HLA-identical donor.

In the past 15 years, several more series havebeen published with encouraging results(Table 1). Transplantation outcomes for pedi-atric patients less than 14 years of age have beenexcellent with reported survival rates now con-sistently[90% although long-term follow-up islimited. Importantly, several studies havedemonstrated that outcomes with 10/10 mat-ched unrelated donors (MUD) are comparableto those with matched sibling donors (MSD)

[66–69]. Tewari et al. reported six patients whoreceived unrelated umbilical cord blood trans-plantation with MAC [70]. All patients survived,and while two of the six patients experiencedgraft failure, they were both successfullyre-transplanted using umbilical cord blood.There have also been a handful of reports ofsuccessful haplo-identical transplantation forCGD in recent years [71–74]. The probability offinding an unaffected MSD in most populationsis less than 25%. The issue of X-linked carriers aspotential MSD is controversial due to increasingevidence of CGD disease burden in carriers thatcan increase with age, as well as the need toensure higher engraftment levels to achievesymptomatic cure than may be needed with anunaffected donor.

Patients with intractable infection or activeinflammation at time of transplantation andadolescents and young adults have remaineddifficult to transplant. Adolescents and youngadults (14 years of age or older) have historicallyhad increased transplant-related mortality ratesbetween 28% and 50% [64–67]. Unfortunately,efforts to reduce toxicity by utilizing reducedintensity conditioning regimens have generallybeen complicated by high rates of graft failure.In 2014, Gungor et al. published a largeprospective, multicenter study that included 56patients aged 0–40 years (median 13 years) whounderwent HCT with RIC using low-dosebusulfan, fludarabine, and serotherapy witheither ATG or alemtuzumab [78]. Importantly,42 of the 56 patients were considered high riskdue to active infection and/or autoinflamma-tion. The group reported an impressive 2-yearoverall survival of 96% and an event-free sur-vival of 91%. The cumulative incidence of gradeIII–IV acute GVHD was low at 4%, and chronicGVHD was 7%. Graft failure occurred in only 3of 56 (5%) patients, and stable (C 90%) myeloidchimerism was found in 93% of survivingpatients. However, subsequent studies havereported high rates of graft failure and mixedchimerism. Khandelwal et al. recently com-pared a cohort of 14 patients who received MACwith busulfan, cyclophosphamide, and ATGversus four patients who received RIC with flu-darabine, melphalan, and alemtuzumab [80].All four patients who received RIC survived, but

2548 Adv Ther (2017) 34:2543–2557

Table1

Transplantation

Outcomes

forCGD

Reference

No.

ofpatients

Patient

agein

years(m

edian)

Don

orsource

Con

dition

ingregimen

GVHD

prop

hylaxis

aGVHD

a

(no.

ofpatients)

cGVHD

(no.

ofpatients)

Overall

survival

Disease-free

survival

Horwitzet

al.

[64]

105–

36(15)

MSD

RIC

:Cy/Flu/ATGandDLI

CSA

12

7/10

(70%

)6/10

(70%

)

Segeret

al.[65]

270.8–

38.7

(8.5)

MSD

(25)

MUD

(2)

MAC

(17):Bu/CyMAC

(1):Bu/Mel/

alem

tuzumab

MAC

(1):

Bu/Cy/ATG

MAC

(1):

Bu/Cy/TNIMAC

(1):

Bu/Cy/TT/A

TG

MAC

(1):Bu/Flu/ATG

RIC

(2):Bu/Flu/ATG

RIC

(1):Flu/Cy/ATG

RIC

(1):Flu/TBI

CSA

(27)

MTX

(13)

prednisone

(4)

43

23/27(85%

)22/27(81%

)

Soncinietal.[66]

201.25–2

1(6.25)

MSD

(9)Sib

UCB(1)

10/10MUD

(8)9/10

MUD

(1)

UCB(1)

MAC

(16):Bu/

Cy±

alem

tuzumab

MAC

(1):Bu/Mel/

alem

tuzumab

RIC

(2):

Flu/Mel/alemtuzumab

RIC

(1):Bu/Flu/

alem

tuzumab

CSA

03

18/20(90%

)18/20(90%

)

Schuetzet

al.

[67]

124–

20(9.5)

MSD

(3)MUD

(9)

MAC(3):Bu/CyMAC(6):

Bu/Cy/

Flu?

alem

tuzumab

orATG

RIC

(2):Flu/Mel/

RIT

RIC

(1):Flu/TBI

Not

reported

02

9/12

(75%

)7/12

(58%

)

Gozdzik

etal.

[68]

61.5–

13(3)

MSD

(2)10/10

MUD

(3)

9/10

MUD

(1)

MAC:Bu/Cy±

ATG

CSA

±MTX

11

6/6(100%)

6/6(100%)

Martinezet

al.

[69]

111–

13(3.8)

MSD

(4)MUD

(7)

MAC:Bu/Cy/Cytarabine

orFlu/alem

tuzumab

MSD

:CSA

?prednisone

MUD:

CSA

?MTX

00

11/11(100%)

11/11(100%)

Tew

ariet

al.[70]

120.67–1

1.6(4.95)

MSD

(5)Sib

CB(1)UCB

(6)

MAC

(5):Bu/Cy±

ATG

MAC

(6):Bu/Flu/Cy/

ATGRIC

(1):Flu/Cy/

ATG

CSA

/MTX

(4)CSA

/MMF(5)CSA

/steroid(2)CSA

/TCD

(1)

14

12/12(100%)

10/12(83.3%

)

Adv Ther (2017) 34:2543–2557 2549

Table1

continued

Reference

No.

ofpatients

Patient

agein

years(m

edian)

Don

orsource

Con

dition

ingregimen

GVHD

prop

hylaxis

aGVHD

a

(no.

ofpatients)

cGVHD

(no.

ofpatients)

Overall

survival

Disease-free

survival

Gun

goret

al.

[78]

560.8–

40(12.7)

MSD

(18)

MRD

(3)

10/10MUD

(25)

9/10

MUD

(10)

RIC

:Bu/Flu?

ATG

oralem

tuzumab

CSA

orTAC?

MMF

24

52/56(93%

)50/56(89%

)

Mehta

etal.[79]

42–

15(10)

10/10MUD(2)

9/10

MUD

(2)

RIC

:Flu/TBI200cG

y/alem

tuzumab

CSA

orTAC?

MMF

11

4/4(100%)

3/4(75%

)

Oshrine

etal.

[81]

31.1–

4(4)

MUD

RIC

:Bu/Flu/alem

tuzumab

Not

reported

00

3/3(100%)

1/3(33%

)

Khand

elwalet

al.

[80]

180.45–1

9.39

(3.18)

MSD

(2)MRD

(1)10/10

MUD

(10)

9/10

MUD

(5)

MAC

(14):Bu/Cy/ATG

RIC

(4):Flu/Mel/

alem

tuzumab

Cy/pred

(15)

Cy/

pred

±Maraviroc

(2)Sirolim

us/pred

(1)

54

15/18(83%

)15/18(83%

)

Morillo-Gutierrez

etal.[73]

703.8–

19.3

(8.9)

MSD

(12)

MRD

(1)

Haplo

(1)

10/10MUD

(44)

9/10

MUD

(11)

UCB(1)

MAC

(46):Treo/

Flu±

ATG

oralem

tuzumab

MAC

(15):

Treo/Flu/TT±

ATG

oralem

tuzumab

Other

(9):

undefin

edtreosulfan

basedregimen

CSA

(4)

CSA

?MMF(44)

CSA

/MMF/pred

(1)CSA

?MTX

(13)

TAC?

MTX

(8)

89

64/70(91.4%

)59/70(84%

)

MSD

Matched

siblingdonor,MRD

matched

relateddonor,MUD

matched

unrelateddonor,UCBum

bilicalcord

blood,Haplohaplo-identicald

onor,M

ACmyeloablative

cond

itioning,

RIC

reducedintensitycond

itioning,Bubusulfan,

Cycyclophosphamide,

Flufludarabine,Mel

melphalan,TT

thiotepa,TBItotalbody

irradiation,

ATG

anti-thymoglobulin

,Treo

treosulfan,

CSA

cyclosporine,T

ACtacrolim

us,M

TXmethotrexate,MMFmycophenolate

mofetil

aGrade

III–IV

aGVHD

2550 Adv Ther (2017) 34:2543–2557

three patients developed mixed chimerism, andtwo required stem cell boosts to maintain donorchimerism. The results at our own institutionusing RIC have also not been successful. Threepatients who underwent HCT with RIC all hadpoor engraftment and needed further inter-vention with either donor lymphocyte infu-sions or repeat HCT [81]. As such, we currentlyuse MAC in all patients with CGD at ourinstitution.

As with all HCT recipients, there is concernregarding the potential for development of lateeffects and durability of immune reconstitutionin CGD patients following HCT. This is of par-ticular concern as many CGD patients may betreated early in life, leading to early exposure tothe toxicities of conditioning agents. Furtherstudy specifically aimed to address late effectsand durability of immune reconstitution areneeded.

Cole et al. found that pediatric patients whounderwent HCT had fewer infections (0.15 epi-sodes of infection/admission/surgery per yearvs. 0.71 episodes infection/admission/surgeryper year) and improved growth parameters(height and BMT) compared to those treatedconventionally [75]. The same group alsodemonstrated that quality of life (QOL) wassignificantly higher in transplanted childrenversus non-transplanted children [76]. In fact,parent-reported and patient-reported QOL intransplanted children were comparable to levelsreported from healthy children. A recentSwedish study directly compared outcomes of14 X-linked CGD patents who underwent HCTwith 13 patients who received conventionalmanagement [77]. Thirteen of 14 (92%) menaged 1–35 years of age survived HCT at medianfollow-up of 7 years (range 1–16) and werecured of disease. Contrarily, 7 of 13 (54%) mentreated conventionally died at a mean age of19 years, and all others suffered fromlife-threatening infections.

GENE THERAPY FOR CGD

Gene therapy is an attractive alternative to HCTand would provide an option for patientswithout an HLA-identical donor. Autologous

HCT also eliminates the risk of GVHD andabrogates the need for long-term immunosup-pressive therapy. Furthermore, full engraftmentis likely unnecessary for CGD, as data fromhealthy female carriers of X-linked CGD indi-cate that as low as 10–20% functional neu-trophils is adequate to protect against severeinfection [53, 54].

The first gene therapy trials for CGD tookplace at the NIH in the 1990s [82]. Five patientswith autosomal recessive p47phox deficiencyreceived gene-corrected CD34 ? hematopoieticstem cells (HSCs) without conditioning using arecombinant c-retroviral vector. Functionallycorrected granulocytes were detected in all 5patients but with a peak of only 0.004–0.05%total circulating granulocytes at 3–6 weeks, andno gene-corrected neutrophils were present at1 year post-gene therapy. Protocol modifica-tions to enhance mobilization of CD34? cellsand to improve retroviral transduction effi-ciency failed to improve long-term engraftment[83]. These initial studies demonstrated thenecessity of at least some degree of conditioningwith gene therapy for CGD.

Several small trials subsequently performedgene therapy for X-linked CGD using c-retrovi-ral vectors and non-myeloablative condition-ing. Two adult men aged 25 and 26 yearsunderwent gene therapy in Germany using ac-retroviral vector expressing gp91phox undercontrol of the spleen focus-forming virus pro-moter and low-dose busulfan at 8 mg/kg[84, 85]. Approximately 15% of peripheralblood neutrophils were found to expressgp91phox within the first 5 months after trans-plantation, and both patients experiencedclinical benefit with resolution of bacterial andfungal infections. However, gene markingincreased with time due to insertional activa-tion of the proto-oncogene MDS1-EVI1, whilemethylation of the viral promoter resulted intransgene silencing and loss of clinical benefit.Both patients ultimately went on to developmyelodysplastic syndrome (MDS); one patientdied at 27 months post-gene therapy from sep-tic shock, and the other patient went on toreceive a MUD HCT at 45 months [85]. Thesame vector and protocol were used to treat twochildren in Switzerland with therapy-resistant

Adv Ther (2017) 34:2543–2557 2551

A. nidulans infections with resolution of infec-tion [86, 87]. However, the same clonal expan-sion was observed in both children, and bothwere rescued with allogeneic HCT [87].

Three patients, aged 28, 28, and 19 years,with X-linked CGD and severe infection notresponsive to conventional managementunderwent gene therapy at the NIH using amurine Moloney retrovirus-derived vector tointroduce gp91phox cDNA into CD34? HSCsand low-dose busulfan at 5 mg/kg/day for 2 days[88]. The team achieved early marking of 26, 5,and 4% of neutrophils, but there was sustainedlong-term marking of only 1.1% and 0.03% ofneutrophils in patients 1 and 3, respectively.Nevertheless, both patients had full or partialresolution of infection. Gene-marked neu-trophils had sustained correction of oxidaseactivity, indicating that silencing of the trans-gene was not a problem with this vector, andgene marking was polyclonal. Unfortunately,patient 2 had no detectable corrected neu-trophils at 4 weeks, and he died at 6 monthspost-gene therapy from a pre-existing fungalinfection.

In total, five phase I/II clinical trials wereperformed in Germany, London, the NIH, andSeoul with 12 patients transplanted usingc-retroviral vectors and RIC [83, 89]. All thetrials demonstrated initial engraftment oftransduced neutrophils at 10–30% of totalneutrophils and clinical benefit with resolutionof pre-existing and life-threatening infections.However, cell engraftment progressivelydecreased with time in all patients, and severalpatients developed MDS. Corrected HSCs do nothave a selective growth advantage compared toNADPH oxidase-deficient cells, contrary to whatis seen with some other primary immunodefi-ciencies. This suggests that more myeloablativeconditioning may be needed for sustainedengraftment. It has also been suggested thatconstitutive expression of gp91phox inhematopoietic progenitor cells may lead to theinappropriate production of ROS with resultanttoxicity and loss of gene-corrected cells overtime.

In response to the high incidence of MDSseen with the c-retroviral vectors and concernsrelated to stem cell toxicity mentioned above,

codon-optimized self-inactivating (SIN) lentivi-ral vectors have been developed wherebytransgene expression is limited to the myeloidlineage. Santilli et al. developed a SIN-lentiviralvector with gp91phox cDNA under the control ofa chimeric promotor that contains binding sitesfor the myeloid transcription factors CAAT boxenhancer-biding family proteins (C/EBPs) andPU.1, which are highly expressed during gran-ulocyte differentiation [90]. This allows for highlevels of gp91phox expression in terminally dif-ferentiated neutrophils while de-targetingexpression in HSC’s. Chiriaco et al. took thisone step further and developed a dual-regulatedlentiviral vector employing a myeloid-specificpromoter and microRNA to post-transcription-ally regulate transgene expression [91]. Preclin-ical trials in mice demonstrated high levels ofgp91phox expression in myeloid cells with spar-ing of the CD34 ? HSC compartment. Multi-center clinical trials using the Santilli et al.vector and low-dose busulfan are currentlyunderway in the United States and Europe. Sofar, three patients have undergone gene therapyin the United States, and early results arepromising, with resolution of CGD phenotypeand sustained neutrophil gene marking withtime (unpublished data).

Finally, targeted genome editing approachesusing zinc-finger nucleases (ZFN), transcriptionactivator-like effector nucleases, or the clusteredregularly interspaced short palindromic repeat(CRISPR)-Cas9 system have been proposed toallow gene correction in situ such that geneexpression remains under the control of thegene’s own cell-specific promoters. However,many CGD patients have unique mutations,and these approaches would require engineer-ing unique systems for each individual patient.Another approach is to target specific ‘‘safeharbors’’ in the genome such as the AAVS1 locus(the common integration site of adeno-associ-ated virus), the disruption of which does notresult in genomic instability. However, theadvantage of gene correction in situ is lost usingthis technique. Thus far, preclinical experi-ments using ZFNs and the CRISPR-Cas9 systemhave demonstrated successful transgeneexpression in CD34? HSCs and inducedpluripotent stem cells from CGD patients

2552 Adv Ther (2017) 34:2543–2557

without off-target effects [92–94]. Nevertheless,gene editing approaches remain limited by lowefficiency and hematopoietic stem cell toxicitythrough the process of transfection, expansion,and selection.

CONCLUSIONS

Life expectancy of CGD patients has increasedmore than three-fold over the last few decadesdue to increased recognition of the disease, theadvent of azole antifungals, and improvedmanagement of infectious and inflammatorycomplications. Nevertheless, the incidence oftriazole-resistant Aspergillus is increasing, andthe management of inflammatory complica-tions remains difficult. Survival following HCThas increased from approximately 85% before2000 to greater than 90% in recent reports, andoutcomes have been encouraging regardless ofthe donor source. Children who undergo HCTare also healthier with better QoL than thosemanaged conservatively. As such, HCT shouldbe considered for all patients with CGDregardless of sex, genetic mutation, and clinicalmanifestations. MAC appears to reduce the riskof graft failure and increase the likelihood oflong-term myeloid engraftment. It is preferableto perform HCT as early as possible, but defini-tive cure can also be considered for adolescentand young adult patients, including those witha history of severe infection and autoinflam-mation. Ultimately, as HCT becomes morewidely available and better tolerated, we expectoverall life expectancy for patients with CGD toincrease substantially over the next severalyears. Furthermore, with the new SIN-lentiviralvectors and optimization of conditioning regi-mens, gene therapy may become a viable alter-native to allogeneic HCT for those without anHLA-matched donor.

ACKNOWLEDGEMENTS

No funding or sponsorship was received for thisstudy or publication of this article. All namedauthors meet the International Committee ofMedical Journal Editors (ICMJE) criteria for

authorship for this manuscript, take responsi-bility for the integrity of the work as a whole,and have given final approval for the version tobe published. During the peer review process,Horizon Pharma was offered an opportunity tocomment on the article. Changes resulting fromcomments received were made by the authorbased on their scientific and editorial merit.

Disclosures. Danielle E. Arnold and JenniferR. Heimall have nothing to disclose.

Compliance with Ethics Guidelines. Thisarticle is based on previously conducted studiesand does not involve any new studies of humanor animal subjects performed by any of theauthors.

Open Access. This article is distributedunder the terms of the Creative CommonsAttribution-NonCommercial 4.0 InternationalLicense (http://creativecommons.org/licenses/by-nc/4.0/), which permits any noncommer-cial use, distribution, and reproduction in anymedium, provided you give appropriate creditto the original author(s) and the source, providea link to the Creative Commons license, andindicate if changes were made.

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