BlueCross and BlueShield of Montana Medical Policy/Codes
Immune Cellular Function Assay to Monitor and Predict Immune Function
Chapter: Medicine: Tests
Current Effective Date: December 27, 2013
Original Effective Date: June 07, 2010
Publish Date: December 27, 2013
Revised Dates: November 28, 2011; February 8, 2012; December 11, 2013
Description

Careful monitoring of lifelong immunosuppression is required to insure long-term viability of solid organ allografts without incurring increased risk of infection.  Monitoring of immunosuppression attempts to balance the dual risks of rejection and infection.  Currently, immunosuppression is determined by testing for clinical toxicity (e.g., leukopenia, renal failure) and by therapeutic drug monitoring (TDM) when available.  However, drug levels are not a surrogate for overall drug distribution or efficacy because pharmacokinetics often differs among individuals due to clinical factors such as underlying diagnosis, age, gender and race; circulating drug levels may not reflect the drug concentration in relevant tissues; and levels of an individual immunosuppressant drug may not reflect the cumulative effect of other concomitant immunosuppressants.  The main value of TDM is the avoidance of toxic levels and monitoring patient compliance.  Further, the appropriate level of immunosuppression may vary from person to person.  Individual immune profiles, such as an immune cell function assay, could support clinical decision-making and help to manage the risk of infection from excess immuno-suppression and the risk of rejection from inadequate immunosuppression in immunosuppressed patients.

ImmuKnow® (Cylex®) is an immune cell function assay cleared for marketing by the United States Food and Drug Administration (FDA) in April 2002 to detect cell-mediated immunity (CMI) in an immunosuppressed patient population.  The assay measures the concentration of adenosine triphosphate (ATP) in whole blood following 15-18 hour incubation with the mitogenic stimulant, phytohemagglutinin (PHA).  In cells that respond to stimulation, increased ATP synthesis occurs during incubation.  Concurrently, whole blood is incubated in the absence of stimulant for the purpose of assessing basal ATP activity.  CD4+ T-lymphocytes are immunoselected from both samples using anti-CD4 monoclonal antibody-coated magnetic particles.  After washing the selected CD4+ cells on a magnet tray, a lysis reagent is added to release intracellular ATP.  A luminescence reagent added to the released ATP produces light measured by a luminometer, which is proportional to the concentration of ATP.  The characterization of the cellular immune response of a specimen is made by comparing the ATP concentration for that specimen to fixed ATP level ranges.

On April 2, 2002, Cylex obtained 510(k) clearance from the FDA to market the Immune Cell Function Assay based on substantial equivalence to two flow cytometry reagents (“predicate devices”) manufactured by Becton Dickinson, the TriTestTM CD4 FITC/CD8 PE/CD3 PerCP Reagent and the MultiTestTM CD3 FITC/CD8 PE/CD45 PerCP/CD4 APC Reagent.  These reagents are used to determine CD4+ T-lymphocyte counts in immunocompromised patients.  The FDA-indicated use of the Cylex Immune Cell Function Assay is for the detection of cell-mediated immunity in an immunosuppressed population.

Policy

Each benefit plan, summary plan description or contract defines which services are covered, which services are excluded, and which services are subject to dollar caps or other limitations, conditions or exclusions.  Members and their providers have the responsibility for consulting the member's benefit plan, summary plan description or contract to determine if there are any exclusions or other benefit limitations applicable to this service or supply.  If there is a discrepancy between a Medical Policy and a member's benefit plan, summary plan description or contract, the benefit plan, summary plan description or contract will govern.

Coverage

Use of an immune cell function assay to monitor and predict immune function after solid organ transplantation maybe considered medically necessary.

Use of an immune cell function assay for all other indications is considered experimental, investigational and unproven.

Rationale

The ImmuKnow® assay has been examined in clinical trials for its potential use in monitoring immunosuppression medication regimens in solid organ transplant patients.

Assessment of a diagnostic technology typically focuses on three analyses:

  • Analytic validity including comparison to a “gold-standard” test and test/re-test reliability;
  • Clinical validity including sensitivity, specificity, positive and negative predictive value in appropriate populations of patients; and
  • Clinical utility that is demonstration that the information from the diagnostic test results in improved health outcomes.

The sensitivity of a test is the ability to detect disease when the disease is present (true positive), while specificity indicates the ability to detect patients who do not have the disease (true negative).  Evaluation of clinical validity, therefore, requires independent assessment by two methods in a population of patients suspected of having a disease but not all of whom do have the disease/disorder.  Additionally, demonstration of the clinical utility of the ImmuKnow assay would require specifying abnormal levels prior to testing an immunosuppressed patient population, making treatment decisions based on the assay results, and documenting decreased morbidity and/or mortality (such as improved transplant organ survival and/or reduced infectious complications) following these treatment decisions.

In support of Cylex’s application for FDA clearance, Kowalski et al. conducted a multi-center study of 155 apparently healthy adults and 127 solid organ transplant recipients (59% kidney, 34% liver, 2% pancreas, 5% simultaneous kidney and pancreas).  Immunosuppressive therapies among the transplant recipients were not limited and included muromonab (lymphocyte-depleting antibody, OKT3), anti-thymocyte globulin, calcineurin inhibitors (cyclosporine, tacrolimus), steroids and mycophenolate mofetil, a purine synthesis inhibitor.  ImmuKnow assays were performed < one month to > four years after transplant.  Additional details on testing were not specified.  Ninety-two percent of the transplant patients had CD4+ ATP levels < 525 ng/ml, while 94% of apparently healthy controls had CD4+ ATP values > 225 ng/ml.  

The authors concluded that this defines three zones of patients’ immune response:

  • Adenosine triphosphate(ATP) level < 225 ng/ml indicates that the patient’s circulating immune cells are showing a low response to phytohemagglutinin(PHA) stimulation and suggests that the patient may be at increased risk of infection;
  • ATP level > 525 ng/ml indicates that the patient’s circulating immune cells are showing a strong response to PHA stimulation and suggests that the patient may be at increased risk of transplant rejection;
  • A moderate ATP level (i.e., between 225 and 525 ng/ml) represents a proposed ideal response to PHA stimulation.

ATP level was not correlated with CD4+ T-cell count.

These transplant recipients were included in a follow-up manufacturer-supported study.  The ImmuKnow assay was completed on 504 immunosuppressed transplant recipients (48% kidney, 30% liver, 17% heart, and 5% small bowel) within 30 days after an episode of infection or rejection.  Because only 5% of patients with ATP levels between 130 ng/ml and 450 ng/ml demonstrated adverse events (either infection or rejection), the authors propose this as the target range for ATP level in immunosuppressed transplant recipients.  Note that this analysis yielded different ATP threshold levels for infection risk and rejection risk than those developed in the earlier study and cited in the product insert.  Further, a 2005 manufacturer-supported study of 37 stable pediatric kidney transplant recipients (mean age = 11.1 years) suggests that in children < 12 years of age, risk intervals are defined by ATP level > 395 ng/ml for rejection and < 175 ng/ml for infection.

A manufacturer-supported single-center study assessed 20 small bowel transplant recipients (70% isolated small bowel, 10% multi visceral, 10% modified multi visceral, 10% simultaneous liver, small bowel and pancreas) undergoing tacrolimus tapering per protocol 60-190 days post-transplant.  Among eight patients successfully tapered from tacrolimus, 70% of ATP levels clustered in the low range (< 225 ng/ml), with 25% of ATP levels occurring in the moderate range and 5% occurring in the strong range.  Incidence of infection was not reported.  Twelve unstable transplant recipients (requiring addition of steroid or OKT3) showed ATP levels with 30% in the low range, 43% in the moderate range, and 27% in the strong range.  This study is often described as using ImmuKnow assay results to guide tacrolimus dosing.  However, adjustments to the tapering protocol were determined primarily by histological examination of biopsy results, which correlated with ATP levels as described for the unstable group.

The relationship between low post-transplant ATP levels (< 225 ng/ml) and recent infection in 57 immunosuppressed adult lung transplant recipients was assessed by Bhorade et al.  One hundred forty-three ImmuKnow assays were performed at routine clinic visits when each patient was on a stable dose of tacrolimus.  Fifteen patients developed infections (bacterial or fungal pneumonia, cytomegalovirus infection); 14 of these (93%) had ATP levels < 225 ng/ml at the time of their infections.  Among the 42 non-infected patients, 16 (38%) had ATP levels < 225 ng/ml.  Without comparing post-infection ATP levels with pre-infection ATP levels, it is not possible to draw conclusions about whether a low ATP level contributes to or results from the development of infection.

Of note in this study was the finding that “African American race was an independent predictor of decreased ImmuKnow assay levels that remained significant with stepwise multiple regression analysis.  Interestingly, African American lung transplant recipients received higher doses of tacrolimus that led to similar tacrolimus trough levels, but had significantly decreased immunoassay levels.”  The authors cite data indicating “African Americans who have undergone renal transplantation require a 37% mean higher dose of tacrolimus to achieve similar blood concentrations” and conclude that African-Americans “may be over-immunosuppressed based on the concurrently obtained ImmuKnow assay.”  Because the authors did not report the incidence of infection among African American patients with ATP level < 225 ng/ml, it is not possible to conclude whether these findings are clinically significant.

Two studies found no correlation between ATP levels as determined by the ImmuKnow assay and outcomes in cardiac transplant recipients.  Rossano et al. studied 83 pediatric patients (median age, 4.9 years) undergoing heart transplant.  ImmuKnow assays were performed at routine follow-up visits from three months to over five years after transplant.  There were 26 episodes of acute rejection, 20 (77%) of which were cell-mediated, and the remainder were humoral rejection.  There were 38 infections.  No difference in ATP levels as measured by ImmuKnow assay was detected between patients with or without acute rejection or with or without infection.  Further, the manufacturer’s reported risk ranges for rejection (ATP level > 525 ng/ml) or infection (ATP level < 225 ng/ml) were not predictive of rejection or infection respectively.  As noted above, however, it may be that pediatric patients’ risks for post-transplant infection and rejection correspond to different ATP levels.

Gupta et al. studied 125 adult heart transplant recipients, the majority of whom underwent ImmuKnow assay testing > one year post-transplant.  There was no apparent correlation between ATP level and rejection (n=3).  For seven patients who developed infection, the median ATP level was 267ng/ml and did not differ from the median ATP level in 104 patients who did not develop infection (282 ng/ml).  There was a significant correlation between ATP level and white blood cell count, but not between ATP level and absolute lymphocyte count, suggesting that non-lymphocytes also may influence the ATP response.  This idea is supported by a 1994 study of CD4+ T-cell responsiveness to three stimulants (including phytohemagglutinin) in HIV+ patients.  The authors suggest that assays performed in clinical laboratories should profile immunoregulatory cytokines (e.g., interleukin-2) which modulate the complex interplay between cellular and humoral immune mechanisms.

The studies cited above report ImmuKnow assay results obtained after transplant.  Reinsmoen and colleagues studied 126 kidney transplant recipients to determine whether pre-transplant immune parameters (ATP level as well as human leukocyte antigen (HLA) mismatch, HLA-specific antibodies, and IFN-gamma precursor frequencies to donor or third-party cells) were associated with post-transplant early acute rejection, unstable creatinine course, and poor graft outcome.  The mean pre-transplant ATP level of recipients who had no clinical reason for a biopsy was significantly different from that of recipients who had biopsy-proven AR at any post transplant time point up to 36 months (285.3 ± 143.2 vs. 414.3 ± 138.5 ng/ml).  Recipients who underwent biopsy but had no diagnosis of acute cellular or antibody-mediated rejection had an intermediate mean value of 333.7 ± 156.3 ng/ml.  Pre-transplant ATP levels were also significantly higher for recipients with early (< 90 days) unstable creatinine levels, a significant predictor of early acute rejection, than for recipients with stable creatinine values (362.8 ± 141.2 vs. 283.4 ± 146.4 ng/ml).  Post-hoc analysis using a cutoff ATP level of 375 ng/ml revealed that recipients with pre-transplant ATP > 375 ng/ml were significantly more likely to experience acute rejection (OR=3.67, 95% confidence interval, 1.195, 11.201).  The immune parameters were not used to guide modifications of the immunosuppression protocol.  Graft survival and incidence of infection was not reported in this study.  In their discussion, Reinsmoen et al. suggest that CD4+ T-cell production of ATP in response to PHA stimulation may depend upon the immunosuppressive medications employed:  “Induction immunosuppression can nonspecifically reduce precursor frequencies; however, not all T cells are depleted equally.  T-cells with a memory phenotype [i.e., CD4+] appear to be less susceptible to depletion than naïve T-cells.  Results from studies of alemtuzumab induction therapy have shown that mature T-cells are profoundly depleted, with memory T-cells, B cells, and monocytes depleted to a lesser degree.  Further, the recipients in these trials often experienced reversible rejection episodes that were characterized by a predominant monocyte, not lymphocyte, infiltrate.  Our results suggest the induction therapy may have had more profound negating effect on the population of cells detected by the ELISPOT [i.e., interferon-gamma producing memory T-cells] but not the CD4+ population of cells detected by the ATP synthesis assay.  The association seen between high pre-transplant ATP levels and acute rejection, as seen in this study, may be immunosuppression protocol dependent.  These results suggest the relevance of the immune parameters studied may be influenced by the immunosuppression protocol employed.”

The American Society of Transplantation (AST) has published recommendations for the screening, monitoring and reporting of infectious complications in immunosuppression trials of organ transplant recipients.  These recommendations define relevant infectious complications to be included in the reporting of immunosuppression trials and recommend specific laboratory monitoring and surveillance methods.  The immune cell function assay is not included in these recommendations.

ClinicalTrials.gov lists five studies assessing the ImmuKnow assay’s ability to predict or monitor the development of infection or rejection in solid organ and hematopoietic stem cell transplant recipients and in multiple sclerosis patients.  None of the studies includes modifications of immunosuppressive medication regimens based on ATP level. 

Without clinical trials demonstrating improved patient outcomes, specifically, reduced incidence of infection, rejection and adverse medication-related effects as a direct consequence of ImmuKnow assay results and subsequent treatment, the evidence is insufficient to permit conclusions concerning the effect of this procedure on health outcomes.  Therefore, the ImmuKnow cell function assay is considered experimental, investigational and unproven.

2011 Update

Israeli et al. studied 50 cardiac transplant recipients.  327 samples were acquired and reviewed.  Results noted that “Longitudinal monitoring of Immuknow levels through serial testing proved to be a reliable method for individual patient immune management.”  The authors concluded “Immunknow assay reliably reflects the cellular immune function of heart transplantation patients, thereby supporting the immune monitoring and management of these patients.  Serial longitudinal Immuknow monitoring allows immune management of therapy according to the individual patient’s immune status.”

Xue et al. identified levels of functional immunity in liver transplant patients as measured by the ImmuKnow assay and addressed its application in monitoring posttransplant infection risk.  Results that the authors reported were for infectious liver transplant recipients.  The average ImmuKnow assay was 128 + or – 84ng/ml.  This assay level was significantly lower (P<0.05) than that found in stable liver transplant recipients (305 + or- 149ng/ml).  The author’s noted in their conclusion that because liver transplant recipients with infection were found to have lower ImmuKnow adenosine triphosphate levels, immune cell functional assay provided a new means of monitoring posttransplant infection.

Kobashigawa et al. studied 296 heart transplant recipients of which 864 immune monitoring assays were obtained.  The results reported by the author noted that during infection, the average immune monitoring score was significantly lower than during steady state.  Conclusions noted by the authors were “The non–invasive immune monitoring test appears to predict infectious risk in heart transplant patients.”

Summary

The above mentioned studies included solid organ transplantation recipients.  These studies were able to demonstrate a relationship between immune monitoring assay levels and the recipients’ immune function.  ImmuKnow assay provides a valuable instrument in the supervision and monitoring of the transplant recipients’ immune condition.  Thus, the coverage statement is changed to indicate use of an immune cell function assay to monitor and predict immune function after solid organ transplantation maybe considered medically necessary.

Coding

Disclaimer for coding information on Medical Policies           

Procedure and diagnosis codes on Medical Policy documents are included only as a general reference tool for each policy.  They may not be all-inclusive.           

The presence or absence of procedure, service, supply, device or diagnosis codes in a Medical Policy document has no relevance for determination of benefit coverage for members or reimbursement for providers.  Only the written coverage position in a medical policy should be used for such determinations.           

Benefit coverage determinations based on written Medical Policy coverage positions must include review of the member’s benefit contract or Summary Plan Description (SPD) for defined coverage vs. non-coverage, benefit exclusions, and benefit limitations such as dollar or duration caps.  

ICD-9 Codes

279.50-279.53, 996.80-996.89, V42.81, V42.84, V42.89, V58.44

ICD-10 Codes

D89.810-D89.813, T86.10 –T86.819; T86.890-T86.899, Z48.24, Z48.280-Z48.288, Z48.290-Z48.298, Z94.0 – Z94.4, Z94.81-Z94.82, Z94.9

Procedural Codes: 86352
References
  1. Shearer GM, Clerici M.  In vitro analysis of cell-mediated immunity: clinical relevance. Clin Chem 1994; 40/11(B):2162-5.
  2. Kowalski R, Post D, Schneider MC et al. Immune cell function testing: an adjunct to therapeutic drug monitoring in transplant patient management.  Clin Transplantation 2003; 17:77-88.
  3. Hooper E, Hawkins DM, Kowalski RJ et al.  Establishing pediatric immune response zones using the Cylex ImmuKnow assay.  Clin Transplant 2005; 19(6):834-9.
  4. Zeevi A, Britz JA, Bentlejewski CA et al. Monitoring immune function during tacrolimus tapering in small bowel transplant recipients.  Transpl Immunol 2005; 15(1):17-24.
  5. Kowalski RJ, Post DR, Mannon RB et al. Assessing relative risks of infection and rejection: a meta-analysis using an immune function assay.  Transplantation 2006; 82(5):663-8.
  6. American Society of Transplantation. Recommendations for Screening, Monitoring and Reporting of Infectious Complications in Immunosuppression Trials in Recipients of Organ Transplantation.  Am J Transplant (2006); 6(2):262-74.  Available at: www.a-s-t.org . (Accessed – 2009 Sept. 9).
  7. Bhorade SM, Janata K, Vigneswaran WT et al. Cylex ImmuKnow assay levels are lower in lung transplant recipients with infection.  J Heart Lung Transplant 2008; 27(9):990-4.
  8. Gupta S, Mitchell JD, Markham DW et al. Utility of the Cylex assay in cardiac transplant recipients.  J Heart Lung Transplant 2008; 27(8):817-22.
  9. Reinsmoen NL, Cornett KM, Kloehn R et al. Pretransplant donor-specific and non-specific immune parameters associated with early acute rejection.  Transplantation 2008; 85(3):462-70.
  10. Rossano JW, Denfield SW, Kim JJ et al. Assessment of the Cylex ImmuKnow cell function assay in pediatric heart transplant patients.  J Heart Lung Transplant 2009; 28(1):26-31.
  11. Immune Cell Function Assay in Solid Organ Tranplantation.  Chicago, Illinois:  Blue Cross Blue Shield Association Medical Policy Reference Manual (2009 August) Medical 2.04.56.
  12. Israeli, M, Ben-Gal, T,Yaari V et al.  Individualized immune monitoring of cardiac transplant recipients by noninvasive longitudinal cellular immunity tests.  Transplantation 2010 Apr27:89(8):968-76. 
  13. Xue F, Zhang J, Han L et al.  Immune cell functional assay in monitoring of adult liver transplantation recipients with infection.  Transplantation 2010 Mar 15:89(5):620-6.
  14. Kobashigawa JA, Kiyosaki KK, Patel JK et al.  Benefit of immune monitoring in heart transplant patients using ATP production in activated lymphocytes.  J Heart Lung Transplant 2010 May; 29(5):504-8.  (Epub 2010 Feb 4)
History
November 2011 Policy Reviewed and updated rationale and references. Policy statement remains investigational
February 2012 Policy updated with literature search; additional investigational indication added for HSCT and all other indications; references 10, 15-25 added; title changed to “Immune Cell Function Assay.”
December 2013 Policy formatting and language revised.  Title changed from "Immune Cell Function Assay in Solid Organ Transplantation" to "Immune Cellular Function Assay to Monitor and Predict Immune Function".  Policy statement previously investigational.  Added the following statement: "Use of an immune cell function assay to monitor and predict immune function after solid organ transplantation maybe considered medically necessary".
BCBSMT Home
®Registered marks of the Blue Cross and Blue Shield Association, an association of independent Blue Cross and Blue Shield Plans. ®LIVE SMART. LIVE HEALTHY. is a registered mark of BCBSMT, an independent licensee of the Blue Cross and Blue Shield Association, serving the residents and businesses of Montana.
CPT codes, descriptions and material only are copyrighted by the American Medical Association. All Rights Reserved. No fee schedules, basic units, relative values or related listings are included in CPT. The AMA assumes no liability for the data contained herein. Applicable FARS/DFARS Restrictions Apply to Government Use. CPT only © American Medical Association.
Immune Cellular Function Assay to Monitor and Predict Immune Function