1. Passenger Lymphocyte Syndrome
Jean Oak MD, PhD
Transfusion Medicine Fellow
Stanford University
Hello and thank you for inviting me to speak. Today I’ll describe a case of passenger lymphocyte syndrome in a liver transplant patient and go over some of the literature findings regarding this rare entity.

2. Case study
71 year old female with cirrhosis secondary to primary biliary cholangitis status post directed liver transplant
Donor type: O neg
Recipient type: AB pos
8 units AB pRBC and 6 units AB FFP intraoperatively
Our case is a 71 year old AB+ female with end stage liver disease secondary to primary biliary cholangitis. She received a liver transplant from a directed donor who is Oneg. The transplant went smoothly, during which received 8U of AB RBC and 6U AB FFP.

3. Acute hemolysis on POD 11 POD 11: 2U AB RBC Hemoglobin Bilirubin 10.1
6.4
6.8
Her initial post-operative course was complicated by a bump in her LFTs and total bilirubin around POD 5. However, around POD8 her Hb, which had been holding steady around 9-10 started trending down on 6.8 by POD11. At that time she received 2units of AB RBCs, and her Hb appropriately bumped to Later that evening, however, she complained of chest pain and a repeat CBC revealed an acute Hg drop of more than 2g/dL, which continued to decrease to 6.4 by POD13. Her bili began to also rise from a baseline of around 2mg/dL to almost 6mg/dL. There was no clinical or radiographic support for bleeding, so the clinical team to decided to evaluate her for hemolysis on POD13.
Hemoglobin
Bilirubin

4. POD13 hemolysis workup LDH: 389 U/L Haptoglobin: 145 mg/dL
Bilirubin (indirect): 5.9 (1.5) mg/dL
Type:
Forward: ABpos
Reverse: O
Screen: negative
DAT: positive
IgG: 2+
C3: w+
Elution: anti-A and anti-B
Hemolysis workup on POD13 revealed a mildly elevated LDH of 389, normal hapto, and increased bilirubin which was suggestive but not definitive of hemolysis. Her reverse type showed development of anti-A and anti-B antibodies and her screen was negative. DAT was positive for both IgG and C3, and the eluated confirmed the presence of anti-A and anti-B on her RBCs.

5. Clinical course POD 13: prednisone 1U O+ 2U O+ 2U O+
Given the timing and acuity of the hemolysis and demonstration of anti-A and anti-B, we made a diagnosis of passenger lymphocyte syndrome. She was started on prednisone and her transfusion support was switched from type specific to Opos. She responded well to the 2U of O+ RBCs and her Hb remained relatively stable and her LDH began trending downwards and returned to within normal limits by POD 21. Hematologist also started her on epo given inadequate reticulocytosis and she was discharged on POD27 and has not required transfusion support since POD24.
POD 13: Started prednisone. Hg goal 7-8g/dL given chest pain
POD16: Hematology consult
POD19: Started erythropoietin qMWF for inadequate reticulocytosis
POD27: Discharged on prednisone taper

6. Passenger Lymphocyte Syndrome (PLS)
Transfer of
donor lymphocytes Y
Donor B cells
recognize
recipient RBCs
1o/2o immune response with
IgG production
DAT+
Hemolysis Y Y Y
The case I’ve just described is pretty classic for PLS. PLS is an underrecognized complication of solid and stem transplants that results in delayed hemolysis. The mechanism is the following: During the transplant there is transfer of donor lymphocytes, which recognizes antigens of the recipient RBCs. This results in a humoral response with IgG and less commonly, IgM production that results in a DAT+ hemolysis
Recipient

7. Clinical manifestations of PLS
Abrupt onset hemolysis 5-15 days after transplant
Median 9 days
Usually mild, self-limited hemolysis lasting ~10 days
Our case lasted 11 days
Usually IgG-mediated extravascular hemolysis
DAT can remain positive for 2-3 weeks (liver) or 5 weeks (kidney)
No influence on overall mortality but significant morbidity
Increased transfusions
Rare cases of AKI or DIC due to massive hemolysis
Clinically PLS is characterized by an abrupt onset of hemolysis 1-2 weeks following a transplant, which is about how long a secondary immune response takes. PLS is usually a mild extravascular hemolysis that lasts around 10 days or so until the immune system adjusts itself but the DAT can remain positive for several more weeks. Since hemolysis generally self-limited, long term followup studies does not show a increase in mortality, but it does cause significant morbidity since these patients often require transfusion support during the hemolytic episode. Massive hemolysis causing kidney injury or DIC have been reported in the literature, but this is very rare.

8. Treatment of PLS No clear regimen showing superior outcome
Supportive treatment
Corticosteroids
RBC transfusions with donor compatible units
Massive hemolysis rare but happens
RBC exchange
Plasma exchange
Rituximab
IVIG
Splenectomy
So how do you manage patients with PLS? Unfortunately there are no large scale studies to help identify a clear treatment regimen. Treatment is predominantly supportive with use of steroids and donor compatible RBC units in most cases. If there is massive hemolysis there may be a need for more aggressive treatment options including red cell or plasma exchange, rituxan, IVIG, or a splenectomy.

9. Donor isohemagglutinin-mediated PLS in solid organ transplant patients
In 1984 there was a landmark study by Ramsey et al in the NEJM showing that isohemagglutinins caused most cases of PLS in liver transplant patients. Metanalysis in 1991 showed that anti-A and anti-B was responsible for the majority of the PLS in other solid organ transplant patients as well, and that patients receiving organs that were rich in lymphocytes such as the liver or heart-lung had a higher rate of PLS. This study showed that anti-A from a O donor was the most common cause of PLS in solid organ transplant patients, and that the A2 phenotype was not protective.
Most common in lymphocyte-rich organ transplants
Anti-A most common
A2 phenotype not protective
Ramsey et al. Transfusion 1991

10. PLS due to non-ABO RBC alloantibodies
Any clinically significant alloantibody
Anti-D most common
Rare cases of anti-c, anti-e, anti-Jk(b), anti-Jk(a), anti-Fya, and anti-K also reported
Donors are usually alloimmunized
Anamnestic response?
But as in the case with hemolytic transfusion reactions, there are a number of non-ABO alloantibodies that can cause PLS. And since the antigenicity isn’t any different whether it’s your lymphocytes responding to transfused RBCs versus the donor lymphocytes responding to your RBCs, it’s not surprising that the antigens that most frequently cause hemolytic transfusion reactions also are the most common ones that cause PLS. PLS due to non-ABO alloAbs are usually due to donors who have already been exposed to these antigens in the past, which suggests that PLS may be a predominantly anamnestic response rather than a primary immune response, which also could help explain the timing of the hemolysis.

11. PLS in liver transplant patients: Review of the literature
Studies show variable alloimmunization/hemolysis rates
Alloimmunization rate ranges from %
Hemolysis rate in ranges %
Antibody specificity
N of cases
Hemolysis (% of cases)
Anti-A 57
43 (75%)
Anti-B 20
11 (55%)
Anti-D 12
12 (100%)
Others (C, Jka, Fya, K, k, M, Kpb) 4
2 (50%)
Total 93
66 (73%)
So what kind of conclusions can we draw regarding PLS? I’ve summarized the data from 93 cases of PLS in liver transplant patients that’s been reported in the literature. Since we don’t have large well controlled studies, we can’t predict the true incidence of alloimmunization. The alloimmunization rate ranges anywhere from around 20 to 60% and around 70% of the cases with serologic evidence of alloimmunization show clinical or laboratory evidence of hemolysis. Not surprisingly, anti-A and anti-B are the most common antibodies in PLS, followed by anti-D.
Romero et al. Blood Transfusion 2015
Solves et al. Transfus Med Hemother. 2015
Gniadek et al. Transfusion 2017

12. Risk factors for PLS No clear demographic risk factor
Minor ABO incompatibility
Unclear if donor isohemagglutinin titer matters
Donors with minor RBC alloantibody
High donor lymphocyte load
Solid organs with high lymphocyte content
Stem cell transplants
nonmyeloablative conditioning regimen
lack of anti-B-cell therapy in GVHD prophylaxis
Peripheral blood stem cell transplant 
So what are the risk factors for developing PLS? A prospective study of around 1000 liver transplant patients from Spain showed no clear demographic risk factor. The data in the literature does reveal some risk factors, mainly minor ABO incompatibility, although it’s unclear whether the isohemagglutinin titer in the donor affects the probability of developing PLS. There is some suggestion that donors with a minor red cell antibody will cause PLS if the recipient has that antigen. And lastly, the more lymphocytes you receive from the donor, the higher the likelihood of PLS. The dosage relationship likely explains why solid organ transplants that have high lymphocyte content has a higher rate of PLS. I didn’t discuss PLS in stem cell transplants in the interest of time, but transferring a lot of lymphocytes or not suppressing B cell activity in SCT patients may also increase the likelihood of PLS.

13. Summary
PLS is an under-recognized cause of delayed hemolysis in patients undergoing solid organ transplants
Abrupt hemolysis 1-2 weeks after transplant
Rate of PLS depends on the type of organ transplanted
Higher lymphocyte transfer -> higher risk
Minor ABO mismatch with an O donor is the most common cause of PLS
Utility of monitoring by DAT
When do you start? How often do you test?
So in summary I’ve hopefully shown you today that PLS is an under-recognized cause of delayed hemolysis in solid organ transplant recipients, especially if there is brisk hemolysis that happens 1-2 weeks after the transplant. The rate of PLS depends on the type of organ, or to be more precise, the dosage of donor lymphocytes that are transplanted. Minor ABO mismatch with an O donor is the most common scenario leading to PLS, which means a routine Ab screen will not pick it up. Therefore some studies suggest routine monitoring of these patients with a DAT, but since we know that around 30% of alloimmunization cases does not show evidence of hemolysis AND we know that DATs remain positive for weeks after a hemolytic episode, this raises the question of how we can best utilize the DAT and whether there are better methods that we could use instead. And with that I thank you for your attention and would be happy to take any questions.

14. Selected References Ramsey et al. Transfusion 1991; 31:77–86
Triulzi et al. Transfusion 1992; 32:829–833
Yazer et al. Curr Opin Hematol. 2007;14(6):664-70
Ramsey et al. NEJM 1984; 311:
Romero et al. Blood Transfusion 2015; 13: 423-8
Solves et al. Transfus Med Hemother. 2015; 42(1):8-14.
Gniadek et al. Transfusion. 2017;57(5):
Salerno et al. Transplantation 1998; 65:261–264