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Vol. 58. Issue 8.
Pages 614-617 (August 2022)
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Vol. 58. Issue 8.
Pages 614-617 (August 2022)
Scientific Letter
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Assessment of Quantiferon®-CMV and Immuknow® Assays in CMV-seropositive Lung Transplant Recipients to Stratify Risk of CMV Infection
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Víctor Monfortea,b,c,
Corresponding author
vmonforte@vhebron.net

Corresponding author.
, Helena Sintesa,b, Piedad Ussettid, Raquel Castejóne, Virginia Luz Pérezf, Rosalía Laportad, Amparo Soleg, José Manuel Cifriánh, Pedro J. Marcosi, Javier Redelj, Ibai Los Arcosb,k, Berta Sáeza,b, Rodrigo Alonsof, Silvia Rosadoe, Juan Escrivag, David Iturbeh, Juan Pablo Ovallei, José Manuel Vaqueroj, Eva Revillaa,b, Alberto Mendozaa,b, Susana Gómez-Ollésa,b,c
a Respiratory Department, Hospital Universitari Vall d’Hebron, Barcelona, Spain
b Department of Medicine, Universitat Autònoma de Barcelona (UAB), Barcelona, Spain
c Centro de Investigación en Red de Enfermedades Respiratorias (CIBERES), Instituto de Salud Carlos III, Madrid, Spain
d Respiratory Department, Hospital Universitario Puerta de Hierro, Madrid, Spain
e Laboratory of Internal Medicine Department, Instituto de Investigación Sanitaria Puerta de Hierro, Madrid, Spain
f Respiratory Department, Hospital Universitario 12 de Octubre, Madrid, Spain
g Lung Transplant Unit, Hospital Universitario y Politécnico la Fe, Valencia, Spain
h Respiratory Department, Hospital Universitario Marqués de Valdecilla, Santander, Spain
i Care Process Management, Respiratory Department and Thoracic Surgery, Área Sanitaria de A Coruña y CEE, A Coruña, Spain
j Respiratory Department, Hospital Universitario Reina Sofía, Córdoba, Spain
k Department of Infectious Diseases, Hospital Universitario Vall d’Hebron, Barcelona, Spain
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To the Director,

Lung transplant recipients (LTR) are at significantly higher risk for cytomegalovirus(CMV) infection than other solid organ transplants (SOT).1 The direct and indirect effect of this infection increases morbidity and mortality in LTR. Universal antiviral prophylaxis for between 6 and 12 months depending on the recipient's serology is recommended.1,2 However, this prolonged use has associated side effects, often requiring prophylaxis discontinuation. Several authors have proposed the Quantiferon®-CMV assay to stratify the risk of CMV disease in SOT, which could help to individualize the duration of antiviral prophylaxis.2–7 However, its role is not well-defined in CMV-seropositive LTR (LTR+)8 who accounts for 80–90% of LTR in Spain. It has been reported that 20% of LTR+ patients with high Quantiferon®-CMV values develop CMV infection.6,8 A plausible explanation for this observation could be that a high immunosuppression inhibits other pathways of the immune response against CMV. Immunosuppression intensity can be measured by Immuknow® assay.9 We hypothesized that the risk of CMV infection in LTR+ could be stratified based on Quantiferon®-CMV and Immuknow® assays.

We studied the risk of significant CMV infection or disease (SICD) between stopping prophylaxis (6 months post-transplant) and study end (12 months post-transplant) according to Quantiferon®-CMV and Immuknow® results at the time of stopping prophylaxis. This was a prospective, observational, multicenter study carried out in 7 centers.

Patients were prospectively enrolled between January 2014 and April 2015. Eligible patients were those ≥18 years old with positive CMV serology pre-transplant who survived for more than 90 days.

Data were collected prospectively and CMV DNA loads were performed during scheduled visits at 3, 6, 7, 8, 9, 10, 11 and 12 months post-transplant. Quantiferon®-CMV and Immuknow® assay were measured at 3, 6, 8, 10 and 12 months post-transplant. Clinicians were blind to assay results.

Quantiferon®-CMV assay (Cellestis, a QIAGEN Company) has been validated to detect CMV-specific immune response by measuring the amount of IFN-γ produced by CD8+ T-cells after ex vivo stimulation with peptides that simulate CMV proteins. Quantiferon®-CMV assay was considered reactive when IFN-γ was ≥0.2IU/mL.3 Indeterminate results (3.4% of assays performed) were not included in the analysis.

The intensity of immunosuppression was assessed by measuring cellular immune function using the Immuknow® assay (Cylex Inc.), which evaluates intracellular ATP levels in CD4+ T-cells. Immuknow® values were low, intermediate or high when ATP was <225, 225–525 or ≥525ng/mL, respectively.10

CMV prophylaxis duration was 6 months. All patients received CMV prophylaxis consisting of intravenous ganciclovir following surgery. Once oral intake was restarted, this was switched to oral valganciclovir 900mg once daily (dose adjusted to renal function) until 6 months post-transplant. Diagnosis of CMV infection/disease was based on established criteria.10 CMV infection was considered significant when CMV DNAemia was higher than 1000copies/mL.

The study was approved by the institutional review board of Hospital Vall d’Hebron (EPA(AG)47/2013). Clinical Trials.gov number NCT02076971.

Seventy-nine (85.9%) patients stopped CMV prophylaxis at 6 months as planned; 13 (14.1%) patients stopped between 3 and 6 months due to adverse reactions.

The proportion of patients free from SCID at 12 months was 78.3% (95% CI, 70.6%–87.4%). No differences were observed between patients who developed SICD and those who did not (Table 1). Twenty-eight cases of SCID were reported in a total of 20 patients (21.7%). No patients developed significant CMV DNAemia while on prophylaxis, and just 2 patients had CMV DNA loads <1000copies/mL, which resolved spontaneously. No patients died of CMV disease. Two patients died due to acute renal failure and septic shock respectively.

Table 1.

Clinical and demographic characteristics of patients who developed significant CMV infection (CMV DNA copies >1000mL–1) or disease and those who did not.

  CMV infection/disease (N=20)  No CMV infection/disease (N=72)  p 
Sex (M/F)  14 (70%)/6 (30%)  42 (58.3%)/30 (41.7%)  0.441 
Age (years)(mean±SD)  57.18±11.32  54.28±11.43  0.317 
Underlying disease      0.384 
Pulmonary fibrosis  11 (55%)  23 (31.94%)   
COPD/emphysema  2 (10%)  22 (30.55%)   
Cystic fibrosis  1 (5%)  9 (12.5%)   
Pulmonary hypertension  5 (25%)  7 (9.72%)   
Other  1 (5%)  11 (15.27%)   
Type of transplant      0.64 
Unilateral  11 (55%)  22 (30.5%)   
Bilateral  9 (45%)  50 (54.34%)   
Immunosuppressive induction      0.211 
No  12 (60%)  31 (43.06%)   
Yes  8 (40%)  41 (56.94%)   
Tacrolimus trough level at 6 months post-Tx (mean±SD)  10.87±2.77  12.82±5.20  0.132 
Donor CMV serology      0.338 
Positive  17 (85%)  49 (68.05%)   
Negative  2 (15%)  14 (19.45%)   
Adverse reactions per patient (mean±SD)  2.29±1.38  1.47±0.61  0.177 
Acute cellular rejection per patient (mean±SD)  1.33±0.71  1.08±0.28  0.150 
Opportunistic infections (mean±SD)  1.62±0.87  2.10±1.17  0.480 
Respiratory function at 6 months post-Tx (mean±SD)
FVC (L)  2.96±1.18  2.67±0.80  0.319 
FEV1 (L)  2.29±0.78  2.02±0.66  0.188 
FEV1/FVC (%)  79.08±11.23  76.95±14.94  0.503 
Lymphocyte total count at 6 months (10*9L–1) (mean±SD)  2.00±0.10  1.97±0.239  0.321 
Glomerular filtration rate (mL/min) at 6 months (mean±SD)  67.75±27.12  75.58±29.79  0.340 

At the first test (3 months post-transplant), Quantiferon®-CMV assay was reactive in 69 (78.4%) patients and non-reactive in 19 (21.6%), with a median IFN-γ of 1.07IU/mL (IQR: 0.29–10.57IU/mL.) At the last test (12 months post-transplant), 73 (84.9%) patients had a reactive assay, and 13 (15.1%) had a non-reactive assay, with a median IFN-γ of 3.87IU/mL (IQR: 0.58–27.49IU/mL). Median Quantiferon®-CMV values increased over time (p=0.001).

At the end of prophylaxis, Quantiferon®-CMV assay was reactive in 70 of 86 patients (81.4%) and non-reactive in 16 (18.6%). The proportion of patients free from SCID at study end was 78.6% (55/70) in the group with reactive Quantiferon®-CMV at the end of prophylaxis and 81.3% (13/16) in the non-reactive group (p=0.872) (Fig. 1).

Fig. 1.

(a) Proportion of patients free from significant CMV infection (CMV DNA copies >1000mL–1) or disease according to Quantiferon®-CMV assay, between the end of prophylaxis and the end of the study. Reactive Quantiferon®-CMV (IFN-γ ≥0.2IU/mL, continuous line) vs. non-reactive (IFN-γ <0.2IU/mL, dashed line). (b) Proportion of patients free from significant CMV infection (CMV DNA copies >1000mL–1) or disease according to Immuknow® assay result from the end of prophylaxis to the end of the study. Intermediate/high Immuknow® value (ATP ≥225ng/mL, continuous line) vs. low value (ATP <225ng/mL, dashed line).

(0.13MB).

At the first test (3 months post-transplant) Immuknow assay was low in 67 (72.8%) patients and intermediate in 25 (27.2%); no patients had high values. The median ATP value was 137.86ng/mL (IQR: 75.50–237.02ng/mL). At the last test (12 months post-transplant) 56 (66.7%) patients had low values, 27 (32.1%) had intermediate values and 1 patient had high values (1.2%), with a median ATP of 177.50ng/mL (IQR: 125.24–254.65ng/mL). Median ATP increased over time (p=0.003).

At the end of prophylaxis, 14 of 91 (15.4%) patients had an intermediate Immuknow® value, 77 (84.6%) had a low value, and none had a high value. The proportion of patients free from SCID at study end was 92.9% (13/14 patients) for those with intermediate values at the end of prophylaxis and 76.6% (59/77 patients) for those with low value (p=0.158) (Fig. 1).

At the end of prophylaxis, no patients had non-reactive Quantiferon®-CMV with intermediate/high ImmuKnow® values. Sixteen (18.8%) patients had non-reactive Quantiferon®-CMV and a low ImmuKnow®. Fifty-six (65.9%) had a reactive Quantiferon®-CMV and a low ImmuKnow®, and 13 (15.3%) had a reactive Quantiferon®-CMV and an intermediate/high ImmuKnow® value. None of these 13 patients developed SICD, compared to 75% of the remaining patients (p=0.056)

The predictive value of both assays was not statistically significant. Quantiferon®-CMV showed an AUC of 0.571 (95% CI, 0.411–0.732; p=0.344) with a sensitivity of 16.7% (95% CI, 4.4%–42.3%) and a specificity of 80.9% (95% CI, 69.2%–89.0%). The positive predictive value (PPV) was 18.8% (95% CI, 5.0%–46.3%) and the negative predictive value (NPV) was 78.6% (95% CI, 66.8%–87.1%). For ImmuKnow®, an AUC of 0.45 (95% CI, 0.29–0.61; p=0.514), a sensitivity of 94.7% (95% CI, 71.9%–99.7%), a specificity of 18.1% (95% CI, 10.3%–29.3%), a PPV of 23.4% (95% CI, 14.8%–34.7%) and a NPV of 92.9% (95% CI, 64.2%–99.6%) were observed.

Quantiferon®-CMV assay has been proposed as a potential tool for predicting CMV infection in SOT after stopping CMV prophylaxis4,6,7,12 and for helping personalize CMV prophylaxis.5 However, most published studies include both seropositive and seronegative patients or/and a small number of lung transplants which makes it difficult to draw conclusions for LTR+. To our knowledge, our study is the first to focus on LTR+ alone. Unlike other SOT, we found no difference in the rate of SCID between patients with reactive and non-reactive Quantiferon®-CMV assays in the months after stopping prophylaxis (around 20% in both groups). Therefore, Quantiferon®-CMV was not useful for risk stratification in this population. Alternate biomarkers such as IFN-γ-CMV ELISPO remain under investigation.13

Little is known about the value of the Immuknow® assay for CMV risk stratification in LTR. Published studies have focused on overall infections in SOT but not specifically CMV infection.11,14–18 Although some studies have suggested its utility for identifying patients at risk of CMV infection in SOT.19,20 In the present study, only 7% of patients with high/intermediate values developed SCID versus 25% of patients with low levels. Therefore Immuknow® could help to individualize the prophylaxis length and CMV DNAemia surveillance. However, differences did not reach statistical significance (p=0.158) and additional studies are needed to confirm this purpose.

Finally, most LT+ had specific immune response to CMV (80% of patients with reactive Quantiferon®-CMV), but the majority were also heavily immunocompromised (70% with low Immuknow® values). This may be the reason why 20% of patients with specific immune response to CMV develop SCID. In fact, no patient with specific immune response and who was not heavily immunosuppressed developed SICD. On the other hand, the observed increase in Quantiferon®-CMV and Immuknow® values over time, probably indicates a certain recovery of immunocompetence.

In conclusion, Quantiferon®-CMV assay was not useful for risk stratification of SCID in LTR+. However, Immuknow® assay may be useful for this purpose, but additional studies are needed to confirm this.

Funding

This work was supported by Roche Farma SA. Madrid, Spain.

Conflict of interest

The authors declare no conflicts of interest.

Acknowledgments

The authors thank Sonia López and Rosa Llòria and for their assistance and technical help and Núria Pajuelo for her work on the statistical analysis.

References
[1]
J. Torre-Cisneros, J.M. Aguado, J.J. Caston, L. Almenar, A. Alonso, S. Cantisán, et al.
Management of cytomegalovirus infection in solid organ transplant recipients: SET/GESITRA-SEIMC/REIPI recommendations.
Transplant Rev, 30 (2016), pp. 119-143
[2]
C.N. Kotton, D. Kumar, A.M. Caliendo, S. Huprikar, S. Chou, L. Danziger-Isakov, et al.
The third international consensus guidelines on the management of cytomegalovirus in solid organ transplantation.
Transplantation, 102 (2018), pp. 900-931
[3]
S. Walker, C. Fazou, T. Crough, R. Holdsworth, P. Kiely, M. Veale, et al.
Ex vivo monitoring of human cytomegalovirus-specific CD8+ T-cell responses using QuantiFERON-CMV.
Transpl Infect Dis, 9 (2007), pp. 165-170
[4]
O. Manuel, S. Husain, D. Kumar, C. Zayas, S. Mawhorter, M.E. Levi, et al.
Assessment of cytomegalovirus-specific cell-mediated immunity for the prediction of cytomegalovirus disease in high-risk solid-organ transplant recipients: a multicenter cohort study.
Clin Infect Dis, 56 (2013), pp. 817-824
[5]
D. Kumar, M. Mian, L. Singer, A. Humar.
An interventional study using cell-mediated immunity to personalize therapy for cytomegalovirus infection after transplantation.
Am J Transplant, 17 (2017), pp. 2468-2473
[6]
G.P. Westall, N.A. Mifsud, T. Kotsimbos.
Linking CMV serostatus to episodes of CMV reactivation following lung transplantation by measuring CMV-specific CD8+ T-cell immunity.
Am J Transplant, 8 (2008), pp. 1749-1754
[7]
L.F. Lisboa, D. Kumar, L.E. Wilson, A. Humar.
Clinical utility of cytomegalovirus cell-mediated immunity in transplant recipients with cytomegalovirus viremia.
Transplantation, 93 (2012), pp. 195-200
[8]
L. Weseslindtner, H. Kerschner, D. Steinacher, R. Nachbagauer, M. Kundi, P. Jaksch, et al.
Prospective analysis of human cytomegalovirus DNAemia and specific CD8+ T cell responses in lung transplant recipients.
Am J Transplant, 12 (2012), pp. 2172-2180
[9]
R. Kowalski, D. Post, M.C. Schneider, J. Britz, J. Thomas, M. Deierhoi, et al.
Immune cell function testing: an adjunct to therapeutic drug monitoring in transplant patient management.
Clin Transpl, 17 (2003), pp. 77-88
[10]
P. Ljungman, M. Boeckh, H.H. Hirsch, F. Josephson, J. Lundgren, G. Nichols, et al.
Definitions of cytomegalovirus infection and disease in transplant patients for use in clinical trials.
Clin Infect Dis, 64 (2016), pp. ciw668
[11]
S.M. Bhorade, K. Janata, W.T. Vigneswaran, C.G. Alex, E.R. Garrity.
Cylex ImmuKnow assay levels are lower in lung transplant recipients with infection.
J Heart Lung Transplant, 27 (2008), pp. 990-994
[12]
G.P. Westall, Y. Cristiano, B.J. Levvey, H. Whitford, M.A. Paraskeva, E. Paul, et al.
A randomized study of quantiferon-CMV-directed versus fixed duration valganciclovir prophylaxis to reduce late CMV following lung trasplantation.
Transplantation, 103 (2019), pp. 1005-1013
[13]
L. Donadeu, E. Revilla-López, M. Jarque, E. Crespo, A. Torija, C. Bravo, et al.
CMV-specific cell-mediated immunity predicts high level of CMV replication after prophylaxis withdrawal in lung transplant recipient.
J Infect Dis, (2020), pp. jiaa727
[14]
S. Husain, K. Raza, J.M. Pilewski, D. Zaldonis, M. Crespo, Y. Toyoda, et al.
Experience with immune monitoring in lung transplant recipients: correlation of low immune function with infection.
Transplantation, 87 (2009), pp. 1852-1857
[15]
M.Y. Shino, S.S. Weigt, R. Saggar, D. Elashoff, A. Derhovanessian, A.L. Gregson, et al.
Usefulness of immune monitoring in lung transplantation using adenosine triphosphate production in activated lymphocytes.
J Heart Lung Transplant, 31 (2012), pp. 996-1002
[16]
M. Takahashi, A. Ohsumi, K. Ohata, T. Kondo, H. Motoyama, K. Hijiya, et al.
Immune function monitoring in lung transplantation using adenosine triphosphate production: time trends and relationship to postoperative infection.
Surg Today, 47 (2017), pp. 762-769
[17]
D. Piloni, S. Magni, T. Oggionni, A. Benazzo, G. Stella, L. Scudeller, et al.
Clinical utility of CD4+ function assessment (ViraCor-IBT ImmuKnow test) in lung recipients.
Transpl Immunol, 37 (2016), pp. 35-39
[18]
V. Monforte, P. Ussetti, R. Castejon, H. Sintes, V.L. Pérez, R. Laporta, et al.
Predictive value of immune cell functional assay for non-cytomegalovirus infection in lung transplant recipients: a multicenter prospective observational study.
Arch Bronconeumol (Engl Ed), (2021),
[19]
J. Torre-Cisneros, J. Fortún, J.M. Aguado, R. de la Cámara, J.M. Cisneros, J. Gavaldá, et al.
Recomendaciones GESITRA-SEIMC y RESITRA sobre prevención y tratamiento de la infección por citomegalovirus en pacientes trasplantados.
Enferm Infecc Microbiol Clin, 23 (2005), pp. 424-437
[20]
M.A. Pérez-Jacoiste Asín, M. Fernández-Ruiz, F. López-Medrano, C. Aquilino, E. González, T. Ruiz-Merlo, et al.
Monitoring of intracellular adenosine triphosphate in CD4+ T cells to predict the occurrence of cytomegalovirus disease in kidney transplant recipients.
Transpl Int, 29 (2016), pp. 1094-1105
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