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The role of vitamin D in pediatric hematopoietic stem cell transplantation

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The role of vitamin D in pediatric hematopoietic stem cell transplantation

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Content THE ROLE OF VITAMIN D IN PEDIATRIC HEMATOPOIETIC STEM CELL
TRANSPLANTATION
by
Rusha Hiren Bhandari
A Thesis Presented to the
FACULTY OF THE USC KECK SCHOOL OF MEDICINE
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
MASTER OF SCIENCE
(CLINICAL, BIOMEDICAL AND TRANSLATIONAL
INVESTIGATIONS)
May 2020
Copyright 2020 Rusha Hiren Bhandari

ii
Table of Contents
List of Tables ................................................................................................................................................ iii

List of Figures ................................................................................................................................... iv

Abstract .......................................................................................................................................................... v

Chapter One: Association Between Vitamin D and Risk for Early and Late Post-Transplant
Complications
Abstract ................................................................................................................................................. 1
Introduction ........................................................................................................................................... 2
Methods ................................................................................................................................................. 3
Patient Population ............................................................................................................................. 3
Statistical Approach .......................................................................................................................... 4
Results ................................................................................................................................................... 5
Vitamin D Monitoring and Supplementation ................................................................................... 6
Transplant-Associated Complications .............................................................................................. 8
Survival ........................................................................................................................................... 14
Discussion ........................................................................................................................................... 16

Chapter Two: Pilot Study of Transplant-Related Events in Patients Receiving Ultra-High-Dose Vitamin
Supplementation
Abstract ............................................................................................................................................... 19
Background ......................................................................................................................................... 20
Objectives ............................................................................................................................................ 23
Study Methods and Procedures ........................................................................................................... 24
Design ............................................................................................................................................. 24
Drug Administration ....................................................................................................................... 25
Planned Specimen and Data Collection ......................................................................................... 26
Outcomes and Endpoints ..................................................................................................................... 29
Statistical Considerations .................................................................................................................... 30
Patient Screening, Eligibility, and Enrollment .................................................................................... 30
Study Population ............................................................................................................................ 30
Inclusion and Exclusion Criteria .................................................................................................... 31
Consent Process .............................................................................................................................. 31
Risk/Benefit Assessment and Alternatives to Participation ................................................................ 32
Safety Monitoring ................................................................................................................................ 33
Privacy and Data Confidentiality ........................................................................................................ 34
Financial Obligations and Funding ..................................................................................................... 34
Current Status of Clinical Trial .......................................................................................................... 35
Enrollment ...................................................................................................................................... 35
Preliminary Results ........................................................................................................................ 35
Adverse Events ............................................................................................................................... 35
Barriers to Enrollment .................................................................................................................... 36
Conclusions ......................................................................................................................................... 36

References ................................................................................................................................................... 38

iii
List of Tables
Table Page
1.1: Patient characteristics and vitamin D supplementation………………………………………………..7
1.2: Follow-up Vitamin D levels……………………………………………….…………………………...8
1.3: Univariate and multivariate analysis of time to GVHD incidence…………………………………...10
1.4: Univariate and multivariate analysis of time to VOD incidence……………………………………..11
1.5: Time to acute GVHD in patients with malignant disease…………………………………………….12
1.6: Time to VOD in patients with malignant disease…………………………………………………….13
1.7: Multivariate analysis of event-free and overall survival……………………………………………...15
1.8: Classification of causes of death per Copelan et al’s scheme and per proximal cause of death……...15
2.1a: Stoss dosing………………………………………………………………………………………….26
2.1b: Toxicity due to vitamin D…………………………………………………………………………...26
2.2: Schedule of Assessments……………………………………………………………………………..28

iv
List of Figures
Figure Page
1.1: Description of the study population……………………………………………………………………6
1.2: Development of acute graft-versus-host disease and veno-occlusive disease according to vitamin D
level…………………………………………………………………………………………………………9
1.3: Overall survival according to baseline vitamin D level………………………………………………14
2.1: Study design…………………………………………………………………………………………..24
2.2: Laboratory monitoring………………………………………………………………………………..25
2.3: Study accrual as of January 2020……………………………………………………………………..35

v
Abstract

Hematopoietic stem cell transplantation (HSCT) is a therapeutic and often curative approach to
pediatric blood disorders and cancers. While HSCT outcomes continue to improve by means of reduced
intensity conditioning regimens and alternative donor sources, complications such as graft-versus-host
disease (GVHD) and veno-occlusive disease (VOD) are often severe and at times fatal. These
complications are mediated by inflammation and endothelial injury.
The vitamin D (VD) receptor is present on all cells. VD has immunomodulatory effects, and may
play a role in mitigating the inflammatory response that is implicated in processes such as GVHD and
VOD. While the relationship between VD and post-HSCT complications requires further investigation,
studies have found an association between VD deficiency and GVHD. Additionally, multiple studies have
found that prior to HSCT, a majority of pediatric patients have VD insufficiency that is not improved with
low-dose supplementation.
We investigated the association between VD and post-HSCT complications, also evaluating
practices of VD monitoring and supplementation (Chapter 1). This was accomplished through a
retrospective study of pediatric patients who received HSCT between January 2013 and April 2018. We
found that the majority of patients were VD insufficient prior to HSCT, and low-dose supplementation
did not significantly increase VD levels. We did not find an association between VD and GVHD or VOD,
but there was a correlation between VD and overall survival.
Building on these findings, we are prospectively investigating an alternative VD dosing regimen,
further characterizing the relationship between VD and the inflammatory response, and evaluating the
relationship between VD and post-HSCT complications. In this ongoing study (Chapter 2), subjects
receive a single oral ultra-high-dose of VD (Stoss) prior to HSCT. Immunophenotyping panels and
cytokine levels are obtained prior to and up to 30 days following Stoss dosing to characterize the change
in immune response with Stoss dosing. VD levels and incidence of GVHD and VOD are monitored for

vi
100 days following Stoss dosing. Preliminary results show that Stoss dosing significantly increases VD
levels prior to HSCT.
These two studies help us to better understand the immunomodulatory role of VD in HSCT and
the efficacy of a regimen such as Stoss dosing in achieving VD sufficiency in pediatric patients receiving
HSCT. Further investigation is needed to determine the effect of VD on post-HSCT complications.

1
Chapter One: Association Between Vitamin D and Risk for Early and Late Post-
Transplant Complications*

In this first chapter, we sought to evaluate whether there was an association between vitamin D
levels and complications or survival following hematopoietic stem cell transplantation. We recognized
that an essential component of this multifactorial question was evaluating our institutional practices of
assessing and supplementing vitamin D during the pre- and post-transplant course. This two-pronged
approach has allowed us to better understand the implications of vitamin D insufficiency in pediatric
hematopoietic stem cell transplantation and has underscored the need for more routine assessment and
higher dosing of vitamin D. Our findings were key in informing our subsequent prospective study aims
and design (Chapter 2).

Abstract
BACKGROUND: Vitamin D (VD) deficiency is a well-described phenomenon in pediatric patients
undergoing hematopoietic stem cell transplantation (HSCT). VD modulates inflammation, and deficiency
pre-HSCT and at day +100 has been associated with graft-versus-host disease (GVHD) and poorer
survival. However, a paucity of data have specifically described the association between VD status and
immune-mediated complications including GVHD and veno-occlusive disease (VOD). Additionally, data
to guide recommendations for VD monitoring and supplementation during HSCT are scarce.
OBJECTIVES: Our primary objective was to evaluate the association between VD and post-HSCT
complications. The key secondary aim was to evaluate the routine use and efficacy of VD monitoring and
supplementation practices. To our knowledge, this is the largest study of its kind in the pediatric
population.
METHODS: This retrospective study evaluated VD level (VDL) before and for one year following
HSCT, VD supplementation practices, and their association with acute GVHD, VOD, and survival in

2
pediatric patients who received autologous and allogeneic HSCT for both malignant and non-malignant
diseases from January 2013 to April 2018.
RESULTS: Of 314 HSCTs, 43% (n=136) had VDL measured prior to HSCT; 61% of this cohort had pre-
HSCT VD insufficiency (<30ng/mL). Neither pre-HSCT nor follow-up VDL were associated with the
incidence of GVHD or VOD. Supplementation did not result in significantly different post-HSCT
VDL.VDL was correlated with overall survival; every 10ng/mL increase in VDL was associated with a
28% decreased risk of death (p=0.01).
CONCLUSIONS: Current accepted VD supplementation regimens for pediatric HSCT do not achieve
sufficient VDL in most patients following HSCT. VD status was associated with all-cause mortality but
not individual comorbidities; prospective studies are required to establish the connection between VD
status, inflammatory-mediated HSCT complications, and potential benefit of VD supplementation prior to
and following HSCT. These studies are needed to inform evidence-based guidelines for monitoring and
supplementing VD during HSCT.
*This work was originally published in Biology of Blood and Marrow Transplantation.
Bhandari R, Malvar J, Sacapano A, Aguayo-Hiraldo P, Jodele S, Orgel E. Association between Vitamin
D and Risk for Early and Late Post-Transplant Complications. Biology of blood and marrow
transplantation : journal of the American Society for Blood and Marrow Transplantation.
2020;26(2):343-350.

Introduction
Vitamin D (VD) is a fat-soluble vitamin necessary for calcium and bone homeostasis, but the VD
receptor is also found nearly ubiquitously on cells throughout the body, including immune cells. The
resulting immunomodulatory effects of VD reduce inflammation and mediate endothelial cell activation
and damage
1,2
. Inflammation due to endothelial cell activation during hematopoietic stem cell transplant
(HSCT) is related to severe HSCT-associated complications such as graft-versus-host disease (GVHD),
veno-occlusive disease (VOD), and transplant-associated thrombotic microangiopathy (TMA)
3
. Immune
dysregulation in the setting of VD deficiency during HSCT may therefore predispose patients to these
transplant-associated complications.

3
Pediatric patients receiving HSCT comprise an at-risk population that is susceptible to VD
deficiency and requires more intensive VD supplementation and monitoring. Decreased exposure to
sunlight, multiple medications potentially affecting VD absorption, compromised gut epithelium, and
mucositis are all factors contributing to particular vulnerability to VD deficiency in pediatric HSCT
patients. Multiple studies therefore investigated patients’ VD status prior to and following HSCT. Up to
70% of pediatric patients are VD insufficient (<30ng/mL) prior to HSCT and through Day +100, many
despite already receiving supplementation
4
. A recent study found that only 3% of pediatric patients with
pre-HSCT VD insufficiency received supplemental therapy, and supplementation did not significantly
change rates of VD insufficiency at day +100
5
. Available data from these small and/or single institution
studies report conflicting results as to whether VD status is associated with post-HSCT survival and
complications such as GVHD
4-6
.
Thus, while prior studies consistently found a majority of patients are VD insufficient prior to
HSCT, it remains controversial whether VD status is associated with survival and GVHD, largely due to a
paucity of data. To our knowledge, the association between VD level (VDL) and non-GVHD transplant-
related complications related to endothelial injury (i.e. VOD and TMA) has also yet to be studied.
Moreover, despite the high rates of VD insufficiency and its potential clinical implications, data for best
practices in VD monitoring and supplementation in pediatric transplant populations remains scarce. The
primary aim of this study was to investigate the association between pre-HSCT VD status, and
inflammation-mediated transplant-related complications including GVHD, VOD, and TMA, primarily
during the first year post-HSCT. A key secondary aim was to examine institutional practices for VD
monitoring and supplementation in our contemporary HSCT cohort.

Methods
Patient Population
We established a retrospective cohort consisting of all patients that received HSCT at our
institution from January 2013 to April 2018. Patients were excluded if they were repeating an allogeneic

4
or autologous HSCT for disease recurrence within one year of initial HSCT, as repeat HSCT within this
short time frame would place patients at a unique risk for transplant complications. However, patients
receiving planned tandem autologous HSCTs were included as a continuous HSCT exposure and
analyzed separately from single autologous HSCT. Following approval by the institutional review board,
we extracted demographic data which included age at time of HSCT, sex, race, and self-reported
ethnicity. Disease-specific data included disease type (classified as malignant versus non-malignant) and
HSCT type (allogeneic, autologous, or tandem). Allogeneic subtypes included related, unrelated, and
haplo-identical HSCT. Cell sources for allogeneic HSCT included bone marrow, peripheral blood stem
cells, and cord blood. Both matched and mismatched sources were included. The use of VD
supplementation, adapted from the Seattle Cancer Care Alliance
7
, and all available VDLs were extracted
during the period of two months prior to admission for HSCT until one year following HSCT. Patients
were classified as VD insufficient (VDL <30ng/ml) or deficient (<20ng/ml); a VDL >50ng/ml was
classified as optimal in accordance with previous studies
8-10
. A VDL was considered baseline if
performed between two months prior to admission for HSCT until up to one week after HSCT. To
evaluate the primary endpoint for HSCT-related complications, data was extracted for acute GVHD
(aGVHD) grade 2-4, chronic GVHD, presence of VOD, presence of TMA, relapse, and survival. GVHD
was graded using standard criteria
11,12
; VOD was diagnosed by providers using modified Seattle or
Baltimore criteria
13,14
. Of note, TMA screening only began in 2017 following the implementation of
current guidelines
15
.
Statistical Approach
The χ
2
test was utilized to compare the distribution of patients for baseline VD assessments and
for each HSCT-related endpoint (i.e. GVHD, VOD, and TMA). GVHD analyses were restricted to
allogeneic HSCT recipients. OS time was defined as the length of time between HSCT and death from
any cause. Relapse was defined as post-HSCT recurrence of malignancy or non-malignant condition.
Event-free survival (EFS) is defined as the minimum time between HSCT and relapse or death. Patients
who remained alive were censored at the time of last contact. Kaplan-Meier survival curves were

5
constructed for each survival and toxicity endpoint. Both log-rank and Wilcoxon tests were utilized to
examine differences in the survival curves with similar results; log-rank test (LRT) results are reported
below. Endpoint-specific Cox regression models analyzed the association between the potential covariates
of age at HSCT, HSCT type, disease type, and VD status with cumulative incidence of GVHD,
cumulative incidence of VOD, EFS, and OS. Confirmatory multivariable analyses were performed using
an unbiased mixed stepwise approach with all candidate predictors from Table 1.1. Unless otherwise
stated, all analyses were performed using 2-sided tests, where a p-value ≤ 0.05 was deemed significant.
All computations were completed using the statistical software Stata version 11
16
.

Results
A total of 396 HSCTs were performed from January 2013 to April 2018. Of these, 314 were
eligible for consideration in this study (Figure 1.1). Patient and transplant characteristics are summarized
in Table 1.1. Patient sex and ethnicity were not associated with a difference in whether baseline VDL was
obtained. Patients who were of black race and at least 10 years of age were more likely to have baseline
VDL measured. Of note, as regular screening for TMA was initiated only in 2017, data for TMA was
sparse (n= 7 events) and precluded analysis as a specific endpoint. Data for isolated grade 2-4 chronic
GVHD was also sparse (n=2 events); thus only grade 2-4 aGVHD was included in the analysis as a
specific endpoint.

6

Figure 1.2. Description of the study population

Vitamin D Monitoring and Supplementation
As shown in Table 1.1, baseline VDLs were obtained in less than half of the cohort (n=136/314,
43%). Of those with baseline VDLs, nearly two-thirds (n= 83/136, 61%) were either insufficient or
deficient prior to HSCT. Less than one in ten patients had optimal VDLs entering HSCT. Patients were
significantly more likely to have baseline VDLs obtained if they were undergoing an allogeneic (65%)
versus autologous (12%) HSCT or entering into a series of tandem HSCT (14%). As shown in Table 1.1,
of those with insufficient or deficient VDLs, only half (n= 42/83, 51%) received VD supplementation.
Neither race nor disease type was associated with whether VD was supplemented. Duration of
supplementation was at least three months in 83% of those with insufficient or deficient levels who
received supplementation. Patients with insufficient or deficient baseline VDLs were significantly more
likely to receive supplementation than those with sufficient baseline VDLs (>30ng/mL) (p=0.01). Of
note, one in ten (n=17/178, 10%) patients without a baseline VDL received supplementation.

7
Baseline Vitamin D
level obtained
p- value Vitamin D
supplemented within
one month of HSCT
p- value
Total
(n=314)
Yes
(n = 136)
Total
(n=136)
Yes
(n=57)

Age at HSCT
1
7.62
(5.89)
9.08
(6.19)
<0.001
2
9.08
(6.19)
9.48
(6.14)
0.529
2

Age Range
3
<10 years 209 74 (35%)
<0.001
4

74 29 (39%)
0.780
4
10-15 years 51 29 (57%) 29 13 (45%)
>15 years 54 33 (61%) 33 15 (45%)
Sex
3
Male 174 78 (45%)
0.546
4
78 29 (37%)
0.195
4

Female 140 58 (41%) 58 28 (48%)
Ethnicity
3
Non-Hispanic 114 46 (40%)
0.247
4

46 20 (43%)
0.917
4
Hispanic 143 69 (48%) 69 29 (42%)
Unknown 57 21 (37%) 21 8 (38%)
Race
3
White 129 47 (36%)
0.028
4
47 20 (43%)
0.896
4

Black 24 16 (66%) 16 8 (50%)
Asian/PI 32 12 (37%) 12 5 (42%)
Other/
Unknown
129 61 (47%) 61 24 (69%)
HSCT Type
3
Allogeneic 182
119
(65%)
< 0.001
4
119 51 (43%)
0.834
4

Autologous 90 11 (12%) 11 4 (36%)
Tandem 42 6 (14%) 6 2 (33%)
Disease Type
3
Non-
malignant
68 39 (57%)
0.008
4
39 20 (51%)
0.160
4

Malignant 246 97 (39%) 97 37 (38%)
Baseline Vitamin D Level
(ng/mL)
1
--
27.38
(13.44)
--
27.38
(13.44)
24.02
(11.71)

Baseline Vitamin
D Level (ng/mL)
5
0-19
--
45 (33%)
--
45 (14%) 23 (31%)
0.083
4

20-29 38 (28%) 38 (12%) 19 (26%)
30-49 41 (30%) 41 (13%) 12 (16%)
50+ 12 (9%) 12 (4%) 3 (4%)
Duration of
Supplementation
5

None 79 0 (0%)
--
<3 months 7 7 (12%)
>3 months 49 49 (86%)
Unknown 1 1 (2%)
1: Arithmetic Mean (standard deviation); 2: t-test; 3: n (row percentage); 4: Chi-squared test; 5: n (column
percentage)
HSCT: hematopoietic stem cell transplant
Table 1.1. Patient characteristics and vitamin D supplementation

8
Of those with baseline VDL assessments, the majority had at least one follow-up VDL obtained
(n=94/136, 69%), with a median time to first follow-up VDL of 2.87 months. There was no significant
difference in median time to follow-up VDL between patients who did and did not receive VD
supplementation (2.78 and 2.94 months, respectively, p=0.97). The mean first follow-up VDL between
patients who did and did not receive VD supplementation were 33.67 and 29.16, respectively (p=0.11)
(Table 1.2). Of note, nearly half of pediatric HSCT patients with sufficient, or even optimal VDLs, prior
to HSCT developed VD insufficiency post-HSCT (n=14/32, 44%). VD insufficiency developed in this
population irrespective of whether routine VD supplementation was administered; VD insufficiency
developed in 7 of 15 (47%) of those with sufficient or optimal baseline VDLs who received
supplementation and in 7 of 17 (41%) in those who did not (p=0.73).

Transplant-Associated Complications (GVHD and VOD)
There was no difference in the incidence of aGVHD or VOD in the cohort for those who were
VD deficient, insufficient, sufficient, or optimal at baseline going into HSCT (LRT p=0.52 for aGVHD,

Vitamin D Supplemented within 1 month of HSCT p-value
No (n = 46) Yes (n = 48) Total (n = 94)

Follow-up vitamin D level (ng/mL)
1
29.16 (14.81) 33.67 (12.39) 31.46 (13.74) 0.112
2

Vitamin D follow-up time (months
since HSCT)
3

2.94 (0.10, 11.50) 2.78 (0.03, 12.02) 2.87 (0.03, 12.02) 0.973
4

Survival Status after HSCT
Alive (n = 65) Deceased (n=29) Total (n = 94)
Vitamin D follow-up time (months
since HSCT)
3

2.99 (0.03, 11.50) 2.63 (0.20, 12.02) 2.87 (0.03, 12.02) 0.392
4

Follow-up vitamin D level (ng/mL)
1
33.28 (14.35) 27.38 (11.47) 31.46 (13.74) 0.054
2

EFS status after HSCT

No event (n = 60)
Relapsed/
Deceased (n=34)
Total (n = 94)
Vitamin D follow-up time (months
since HSCT)
3

3.09 (0.03, 11.50) 2.69 (0.10, 12.02) 2.87 (0.03, 12.02) 0.172
4

Follow-up vitamin D level (ng/mL)
1

33.82 (14.21) 27.29 (11.98) 31.46 (13.74) 0.026
2

1: Arithmetic Mean (SD); 2: t-test; 3: Median (Range); 4: Wilcoxon Rank Sum test
† Last available vitamin D level at follow-up. Not all follow-up vitamin D levels occurred prior to relapse.
HSCT: hematopoietic stem cell transplant; EFS: event-free survival
Table 1.2. Follow-up Vitamin D levels†

9
LRT p=0.84 for VOD) or at follow up (LRT p=0.71 for aGVHD, LRT p=0.27 for VOD). Mean VDL at
follow-up for those with aGVHD versus those without were 31.85 ng/mL and 31.42 ng/mL, respectively
(p=0.91). Mean VDL at follow-up for those with VOD versus those without were 30.31 ng/mL and 31.90
ng/mL, respectively (p=0.62) (Figure 1.2). In multivariate analyses, only malignant diagnosis was
associated with incidence of aGVHD (hazard ratio [HR] = 3.50, 95% confidence interval [95%CI] 1.18,
10.36, p=0.01); only autologous HSCT was associated with incidence of VOD (HR= 0.27, 95%CI 0.06,
1.16, p=0.03) (Tables 1.3 and 1.4). In subset analyses of patients with malignant diagnoses, vitamin D
was not associated with risk of aGVHD or VOD (Tables 1.5 and 1.6).

Figure 1.2. Development of acute graft-versus-host disease (a&b) and veno-occlusive disease (c&d)
according to vitamin D level

10
aGVHD
1
Univariate Analysis Multivariate Analysis
Total N %
HR
(SE)
95% CI
HR
p-
value
HR
(SE)
95% CI
HR
p-
value
Gender Male 71 19 70.4 1 --
0.138 --
Female 48 8 29.6
0.57
(0.24)
(0.25,
1.3)
Age at HSCT <10 years 64 14 51.9 1 --
0.996
1 --
0.636
10-15 years 26 6 22.2
1
(0.49)
(0.38,
2.6)
0.72
(0.36)
(0.27,
1.93)
>15 years 29 7 25.9
1.04
(0.48)
(0.42,
2.58)
0.64
(0.32)
(0.24,
1.70)
Race/Ethnicity White/
Non-
Hispanic
24 5 18.5 1 --

0.244
--
Hispanic 62 12 44.4
0.83
(0.44)
(0.29,
2.37)
Black/
Non-
Hispanic
13 2 7.4
0.65
(0.54)
(0.13,
3.34)
Others/
Unknown
20 8 29.6
1.96
(1.12)
(0.64,
5.99)
Is diagnosis
malignant?
Non-
malignant
39 4 14.8 1 --
0.036
1 --
0.011
Malignant 80 23 85.2
3.11
(1.69)
(1.08,
9.01)
3.50
(1.93)
(1.18,
10.36)
Per 10ng/mL Increase in Baseline Vitamin D Level

1.03
(0.14)
(0.78,
1.35)
0.86
Baseline Vitamin
D Level
0-29 ng/mL 73 19 70.4 1 --
0.383
1 --
0.226
>30 ng/mL 46 8 29.6
0.69
(0.29)
(0.3,
1.58)
0.59
(0.26)
(0.24,
1.42)
Vitamin D
Supplemented?
No 68 17 63.0 1 --
0.555 --
Yes 51 10 37.0
0.79
(0.32)
(0.36,
1.73)
Duration Vitamin
D supplemented
after HSCT
None 68 17 63.0 1 --

0.537

--
<3 months 6 2 7.4
1.56
(1.17)
(0.36,
6.79)
>3 months 45 8 29.6
0.7
(0.3)
(0.3,
1.63)
1: n (column percentage)
HSCT: hematopoietic stem cell transplant; aGVHD: acute graft-versus-host disease; HR: hazard ratio; SE: standard
error; CI: confidence interval
Table 1.3. Univariate and multivariate analysis of time to GVHD incidence

11
VOD
1
Univariate Analysis Multivariate Analysis
Total N % HR
(SE)
95%
CI
HR
p-
value
HR
(SE)
95%
CI
HR
p-
value
Gender Male 78 24 66.7 1 -- 0.204 --
Female 58 12 33.3 0.64
(0.23)
(0.32,
1.28)
Age at HSCT <10 years 74 23 63.9 1 -- 0.418 1 -- 0.297
10-15 years 29 7 19.4 0.74
(0.32)
(0.32,
1.72)
0.65
(0.29)
(0.27,
1.56)
>15 years 33 6 16.7 0.56
(0.26)
(0.23,
1.39)
0.50
(0.24)
(0.19,
1.31)
Race/ Ethnicity White/
Non-
Hispanic
28 12 33.3 1 -- 0.05 --
Hispanic 69 19 52.8 0.53
(0.2)
(0.26,
1.09)
Black/
Non-
Hispanic
15 2 5.6 0.23
(0.17)
(0.05,
1.02)
Others/
Unknown
24 3 8.3 0.24
(0.15)
(0.07,
0.84)
HSCT type Allogeneic 119 34 94.4 1 -- 0.161 1 -- 0.035
Autologous 17 2 5.6 0.56
(0.41)
(0.13,
2.34)
0.27
(0.20)
(0.06,
1.16)
Is diagnosis
malignant?
Non-
malignant
39 8 22.2 1 -- 0.28 1 -- 0.060
Malignant 97 28 77.8 1.54
(0.62)
(0.7,
3.38)
2.09
(0.86)
(0.93,
4.70)
Per 10 ng/mL Increase in Baseline Vitamin D
Level
1.11
(0.14)
(0.87,
1.42)
0.38 --
Baseline Vitamin
D Level
0 - 29
ng/mL
83 20 55.6 1 -- 0.361 1 -- 0.680
30+ ng/mL 53 16 44.4 1.36
(0.46)
(0.7,
2.62)
1.16
(0.41)
(0.58,
2.34)
Vitamin D
Supplemented?
No 79 23 63.9 1 -- 0.449 --
Yes 57 13 36.1 0.77
(0.27)
(0.39,
1.52)
Duration Vitamin
D supplemented
after HSCT
None 79 23 63.9 1 -- 0.89 --
<3 months 7 2 5.6 1.02
(0.75)
(0.24,
4.34)
>3 months 49 11 30.6 0.75
(0.28)
(0.37,
1.54)
1: n (column percentage)
HSCT: hematopoietic stem cell transplant; VOD: veno-occlusive disease; HR: hazard ratio; SE:
standard error;
CI: confidence interval
Table 1.4. Univariate and multivariate analysis of time to VOD incidence

12
With
GVHD
Univariate Analysis Multivariate Analysis
Total N % HR
(SE)
95%
CI HR
p-
value
HR
(SE)
95%
CI
HR
p-
value
Gender Male 44 16 69.6 1 -- 0.094 --
Female 36 7 30.4 0.47
(0.21)
(0.19,
1.14)
Age at HSCT <10 years 36 10 43.5 1 -- 0.985 1 -- 0.887
10-15 years 20 6 26.1 1.02
(0.53)
(0.37,
2.8)
0.92
(0.49)
(0.33,
2.59)
>15 years 24 7 30.4 0.93
(0.46)
(0.35,
2.45)
0.77
(0.41)
(0.27,
2.19)
Race/ Ethnicity White/Non-
Hispanic
15 4 17.4 1 -- 0.237 --
Hispanic 50 12 52.2 0.74
(0.43)
(0.24,
2.32)
Black/Non-
Hispanic
4 1 4.4 0.86
(0.96)
(0.1,
7.69)
Others/
Unknown
11 6 26.1 2.08
(1.35)
(0.59,
7.4)
Per 10ng/mL increase in Baseline Vitamin D Level 1.03
(0.16)
(0.76,
1.4)
0.846 --
Baseline
Vitamin D Level
0 - 29
ng/mL
49 16 69.6 1 -- 0.45 1 -- 0.370
30+ ng/mL 31 7 30.4 0.71
(0.32)
(0.29,
1.73)
0.65
(0.32)
(0.25,
1.69)
Vitamin D
Supplemented?
No 49 15 65.2 1 -- 0.771 --
Yes 31 8 34.8 0.88
(0.39)
(0.37,
2.08)
Duration
Vitamin D
supplemented
after HSCT
None 49 15 65.2 1 -- 0.587 --
<3 months 2 1 4.4 2.42
(2.53)
(0.31,
18.74)
>3 months 29 7 30.4 0.81
(0.37)
(0.33,
1.98)
Table 1.5. Time to acute GVHD in patients with malignant disease

13
With
VOD
Univariate Analysis Multivariate Analysis
Total N % HR
(SE)
95%
CI
HR
p-
value
HR
(SE)
95%
CI
HR
p-
value
Gender Male 51 18 64.3 1 -- 0.139 NA
Female 46 10 35.7 0.56
(0.22)
(0.26,
1.21)
Age at HSCT <10 years 46 16 57.1 1 -- 0.52 1 -- 0.592
10-15 years 23 6 21.4 0.74
(0.36)
(0.29,
1.9)
0.72
(0.35)
(0.27,
1.87)
>15 years 28 6 21.4 0.59
(0.28)
(0.23,
1.51)
(0.62
(0.32)
(0.22,
1.69)
Race/Ethnicity White/Non-
Hispanic
19 10 35.7 1 -- 0.075 --
Hispanic 57 16 57.1 0.42
(0.17)
(0.19,
0.93)
Black/Non-
Hispanic
6 0 0
Others/
Unknown
15 2 7.1 0.19
(0.15)
(0.04,
0.87)
HSCT type Allogeneic 80 26 92.9 1 -- 0.112 1 -- 0.042
Autologous 17 2 7.1 0.31
(0.23)
(0.07,
1.31)
0.28
(0.21)
(0.07,
1.21)
Per 10ng/mL increase in
Baseline Vitamin D Level
1.2
(0.18)
(0.9,
1.6)
0.217 --
Baseline Vitamin
D Level
0 - 29
ng/mL
59 15 53.6 1 -- 0.299 1 -- 0.050
30+ ng/mL 38 13 46.4 1.36
(0.46)
(0.7,
2.62)
1.32
(0.54)
(0.59,
2.93)
Vitamin D
Supplemented?
No 60 20 71.4 1 -- 0.239 --
Yes 37 8 28.6 0.77
(0.27)
(0.39,
1.52)
Duration
Vitamin D
supplemented
after HSCT
None 60 20 71.4 1 -- 0.721 --
<3 months 3 1 3.6 1.02
(0.75)
(0.24,
4.34)
>3 months 33 7 25 0.75
(0.28)
(0.37,
1.54)
Table 1.6. Time to VOD in patients with malignant disease

14
Event-Free and Overall Survival
There was no significant difference in OS for those who were VD deficient, insufficient,
sufficient, or optimal at baseline prior to HSCT (p=0.51) (Figure 1.3). Patients who proceeded to
experience relapse or death had significantly lower mean VDLs at follow-up than those who did not
(27.29 ng/mL vs 33.82 ng/mL, respectively; p=0.03). In multivariate analysis, however, only malignant
diagnosis was associated with decreased EFS (p<0.01) (Table 1.7). However, a significant difference in
mean VDL at follow-up was found for patients who did not survive compared to those who did (27.38
ng/mL vs 33.28 ng/mL, respectively; p=0.05). This was confirmed on multivariate analysis for OS, where
every 10ng/mL increase in VDL was associated with a 28% decrease in the risk of death (p=0.01) (Table
1.7). Causes of death per Copelan et al’s algorithm
17
and proximal causes of death are detailed in Table
1.8. Of note, GVHD, VOD, and TMA contributed to a quarter (24%) of the proximal causes of death.
Post-transplant infectious mortality was the single largest proximal cause of death (n= 12/42, 29%).

Figure 1.3. Overall survival according to baseline vitamin D level

15

Total number of deaths 42
Cause of death per Copelan et al’s scheme
17

Primary disease recurrence or
progression
18 (43%)
Organ failure^ 9 (21%)
Infection 7 (17%)
Graft-versus-host disease 4 (10%)
Other* 2 (5%)
Hemorrhage 1 (2%)
Unknown 1 (2%)
Proximal cause of death
Sepsis/infection 12 (29%)
Graft-versus-host disease
+
8 (19%)
Cardiac or respiratory failure 7 (17%)
Unknown 6 (14%)
Intracranial hemorrhage, stroke, or
herniation
4 (10%)
Secondary malignancy 2 (5%)
Veno-occlusive disease
#
1 (2%)
Thrombotic microangiopathy 1 (2%)
Pulmonary hemorrhage 1 (2%)
^1 patient had veno-occlusive disease as cause of death
*1 patient had thrombotic microangiopathy as cause of death
+
3 of these patients also had respiratory failure; 3 had
infectious complications; 1 had multi-organ failure; 1 had
renal failure and died of sudden cardiac arrest
#
This patient also had pulmonary toxicity
Table 1.8. Classification of causes of death per Copelan et al’s
scheme
17
and per proximal cause of death
Event-free survival Overall Survival
Event HR
(SE)
95% CI
HR
p-
value†
Deceased HR
(SE)
95%
CI HR
p-
value†
Diagnosis
Type
Non-
malignant
6
(11.5%)
1 --
<0.001
6
(14.3%)
1 --
0.009
Malignant
46
(88.5%)
3.59
(1.69)
(1.51,
8.54)
36
(85.7%)
2.85
(1.27)
(1.19,
6.86)
HSCT type
Allogeneic
43
(82.7%)
1 --
0.8699
38
(90.5%)
1 --
0.090
Autologous
6
(11.5%)
1.26
(0.56)
(0.53,
3.02)
2
(4.8%)
0.26
(0.19)
(0.06,
1.12)
Tandem
3
(5.8%)
1.10
(0.66)
(0.34,
3.58)
2
(4.8%)
1.00
(0.74)
(0.23,
4.24)
Vitamin D
level, time
dependent
Per 10
ng/mL
increase
--
0.99
(0.11)
(0.90,
1.22)
0.9217 --
0.72
(0.10)
(0.54,
0.94)
0.012
HSCT: hematopoietic stem cell transplant; HR: hazard ratio; SE: standard error; CI: confidence interval
† P-value from a Likelihood ratio test
Vitamin D level treated as a time dependent variable and includes the baseline and follow-up levels
No interactions exist between baseline vitamin D and follow-up levels, p = 0.3054
No interactions exist between follow-up Vitamin D level and transplant type, p = 0.5166
Table 1.7. Multivariate analysis of event-free and overall survival

16
Discussion
In our cohort, we found that less than half of patients were checked for VD insufficiency pre-
HSCT, only half of those who were checked and found to be insufficient actually received VD
supplementation, and current supplementation regimens did not significantly increase VDLs. We also
found that while baseline VD was not associated with relapse, aGVHD, or VOD, higher VDL was
significantly associated with non-relapse, all-cause mortality. Our findings therefore suggest a
relationship between VD and long-term outcomes, and identify opportunities for improvement of VD
monitoring and supplementation.
Few pediatric studies have reported the frequency with which VDL is monitored and
supplemented in HSCT populations. In a study of adult HSCT patients, Sproat et al found that only 20%
of patients had VDL assessed after HSCT, 90% of whom had a low VDL
18
. Earlier studies have raised
similar concerns regarding poor efficacy of routine VD supplementation in HSCT. In a study of adult
patients by Caballero-Velazquez et al, those who received no or low-dose VD (1,000IU per day)
remained insufficient by day +21, as compared to patients receiving 5,000IU VD per day beginning five
days before HSCT who achieved significantly higher serum VDLs by day +14
19
. Wallace et al
investigated standard-dosed VD therapy per National Kidney Foundation guidelines (up to 8,000 IU/day
or 50,000 IU/week) compared to high-dose VD (target dose >200 IU/kg/day). With the standard dosing,
only 43% of patients achieved sufficient VDLs. With the higher dosing, still only 64% of patients
achieved sufficient VDLs
20
. Nonetheless, as demonstrated in our contemporary cohort, routine low-dose
supplementation continued to be used with continued poor efficacy for VD repletion. Our findings
support that patients receiving HSCT are already at high risk for VD insufficiency and require more
aggressive VD supplementation than routine low-dose daily or weekly VD dosing. Single ultra-high-dose
VD (Stoss) is an alternative weight- and VDL-based dosing regimen of 7,000-14,000 IU/kg (maximum
600,000 IU) demonstrating preliminary safety and efficacy in a single-institution study of pediatric
patients receiving HSCT
21
. Further study is necessary to determine if Stoss-dosed VD supplementation
will be effective over large, diverse populations and HSCT regimens. Therefore, we are currently

17
prospectively studying the efficacy and tolerability of Stoss-dosed VD supplementation across diverse
patient populations and conditions.
Our data did not find a significant difference in the cumulative incidence of VOD or aGVHD in
patients according to baseline or follow-up VDL. This is contrary to previous studies in adults and smaller
studies in pediatrics of VD in HSCT. It is possible, as our findings suggest, that there is no association
between VD and GVHD or VOD. However, the pathophysiology of VD insufficiency and inflammation
continue to implicate VD as a regulator of inflammatory-mediated HSCT complications. Studies of the
immunomodulatory effects of VD have found that VD inhibits proliferation of T helper (Th) 1
lymphocytes and their cytokines, indirectly leading to development of Th 2 cells and their cytokines. Th1
cells are the primary mediators of graft rejection, while Th2 cells have a regulatory role
1
. VD has also
been shown to inhibit lipopolysaccharide-induced cytokine production in monocytes and macrophages
22
.
Cytokine production can lead to endothelial cell activation and damage throughout the body, leading to
HSCT complications including GVHD, VOD, and TMA
3
. We therefore postulate that the absence of an
association in our cohort may have been instead due to the high prevalence of VD insufficiency, the
substantial delay in monitoring of post-HSCT VDLs, or the lower overall rate of aGVHD in our cohort
(23%) compared to many of the previous studies. The median time to follow-up VDL was greater than
two months, thereby leaving VD status unknown. This surpasses much of the critical post-transplant acute
period during which most severe transplant complications occur and during which patients likely would
derive the most benefit from optimal VD status. A prospective study with more aggressive VD
supplementation and closer monitoring of VDLs may better elucidate whether VD status is associated
with the incidence of these complications.
Despite the absence of an association with specific transplant comorbidities, we did find a
significant reduction in risk of all-cause mortality with increasing pre-HSCT VDL. While the etiology of
this connection is unclear, immune and inflammatory conditions such as infection, GVHD, VOD, and
TMA contributed to over half of the causes of non-relapse mortality. This further suggests the
immunomodulatory effects of VD may indeed play a key role in post-HSCT survival and that VD acts

18
indirectly to mitigate some of the morbidity from GVHD, VOD, TMA, and infections. We hypothesize
that ongoing cytokine-mediated injury, which is not routinely assessed in HSCT recipients, may affect the
severity of these inflammatory-mediated complications and increase susceptibility to additional insults
such as infection. Studies in critically ill children have also found an association between VD deficiency
and greater illness severity, multi-organ dysfunction, and higher rates of infection
23
. Alternatively, VDL
may instead be serving as a surrogate indicator for overall health status and thus underlying susceptibility
to transplant complications. These hypotheses are being explored in the context of our prospective trial
for Stoss-dosed VD to better understand the connection between VD, VD-mediated changes in cytokines
and inflammation, and HSCT complications and survival.
The limitations of our study are primarily inherent to its retrospective nature. Substantial
variability was present in timing for baseline and follow-up VDLs, thus precluding a direct and consistent
comparison across time points and patients. Similarly, because not all follow-up VDLs were obtained
prior to the diagnosis of GVHD or VOD, VD status prior to the event was interpolated but we were
unable to characterize the temporal distance between VDLs and these complications. As a retrospective
study, undetectable selection bias may be present in terms of which patients had VDL monitored and
supplemented. Lastly, as routine supplementation, patient compliance with the prescribed VD regimen
was often not documented and could not be confirmed. Nonetheless, to our knowledge, this represents the
largest pediatric cohort to date describing VD practices in pediatric patients receiving HSCT. Children
undergoing HSCT clearly comprise an at-risk population that is susceptible to VD insufficiency and
requires intensive VD monitoring and dose-intensive supplementation. Rigorous prospective studies such
as those underway are essential to definitively understand the association between VD supplementation,
VDL, inflammation, and HSCT outcomes. Meanwhile, with VD’s favorable safety profile and potential
health benefit, we propose it would be prudent to routinely monitor VD status prior to and throughout the
first 100 days post-HSCT process with a goal to maintain VD sufficiency throughout.

19
Chapter Two: Pilot Study of Transplant-Related Events in Patients Receiving Ultra-High-
Dose Vitamin Supplementation

In chapter 1, we described an association between vitamin D insufficiency and overall survival,
although the exact mechanism is unclear. We also confirmed that in the transplant population, routine
low-dose vitamin D supplementation did not reliably achieve vitamin D sufficiency. With these findings
in mind, our next step was to evaluate the efficacy of a higher dose vitamin D regimen in achieving and
maintaining vitamin D sufficiency in the immediate post-transplant period. This enabled us to more
clearly evaluate the association between vitamin D and post-transplant complications.
We also sought to investigate the immunomodulatory effects of vitamin D. In vitro, vitamin D
downregulates the pro-inflammatory response. This offers a compelling explanation for why vitamin D
could mitigate transplant complications related to inflammation. We thus obtained cytokine and
immunophenotyping panels to understand the in vivo response to vitamin D and how this correlates with
transplant-related complications. Here we present our study protocol and preliminary results.

Abstract
Studies have found a high prevalence of vitamin D deficiency (up to 70%) in patients prior to
hematopoietic stem cell transplant (HSCT)
4
. Patients with sufficient Vitamin D levels >30ng/mL) prior
to allogeneic transplant have significantly better overall survival (OS) and lower rates of rejection and
relapse
6
. Vitamin D inhibits T helper (Th) 1 and augments Th2 development
1
. Patients who receive
vitamin D supplementation during allogeneic transplant have less inflammatory-mediated processes such
as chronic graft versus host disease (GVHD) and lower levels of naïve CD8+ cells and CD40 ligand
19
.
Multiple studies have raised concern regarding the adequacy of standard and high-dose vitamin D dosing
for vitamin D deficiency. A single oral ultra-high dose of vitamin D given prior to HSCT has been shown
to be a safe and well tolerated method of sustaining therapeutic vitamin D levels for 6-19 weeks
21
.

20
This is a pilot study to investigate whether the single ultra-high-dose regimen is more effective
than daily or weekly dosing and to compare its impact on the incidence of transplant-related
complications, specifically those related to endothelial injury, for 100 days following transplant. We will
also evaluate the dynamic changes in inflammatory biomarkers following ultra-high-dose vitamin D
supplementation. The study population is patients with total vitamin D level <50ng/mL prior to HSCT.

Background
Vitamin D is involved in calcium and bone homeostasis and has immunomodulatory effects.
Vitamin D deficiency prior to HSCT has been associated with a higher risk of acute and chronic GVHD
and inferior survival in adults and children
1,4,6,19,24,25
. Ideal serum concentrations of vitamin D have not
been defined, but levels >50ng/mL would be considered optimal and may be associated with better
outcomes following HSCT
6,9
. Studies have found a high prevalence of vitamin D deficiency in patients
prior to hematopoietic stem cell transplant (HSCT)
4,26
. A prospective study at Cincinnati Children’s
Hospital Medical Center found that despite regular supplementation of vitamin D, 70% of patients had
vitamin D level <30ng/mL prior to HSCT. Among patients who survived 100 days post-transplant, 76%
of those who were deficient prior to HSCT had persistence of the deficiency. Furthermore, 57% of
patients who had normal vitamin D levels before HSCT developed vitamin D deficiency by day 100
4
.
Preliminary institutional data here at Children’s Hospital Los Angeles (CHLA) has shown that a majority
of patients (>90%) who have vitamin D levels checked prior to HSCT have a level <50ng/mL.
27
Vitamin
D deficiency has been associated with greater illness severity in other patients as well. In a meta-analysis
of patients admitted to the pediatric intensive care unit, Vitamin D deficiency was associated with greater
illness severity, multiple organ dysfunction, and mortality
23
. While these studies did not specifically
evaluate patients undergoing HSCT, these associations can be extrapolated and applied to critically ill
HSCT patients.

21
Vitamin D also has immunomodulatory effects. The vitamin D receptor is present in various cells
of the immune system. In vitro, vitamin D inhibits T lymphocyte proliferation, thus inhibiting
transcription of several Th1 lymphocyte cytokines, including interferon (IFN)-gamma and interleukin
(IL)-2. The inhibition of IL-2 expression prevents further T lymphocyte proliferation. This indirectly
leads to T lymphocytes developing into Th2 cells and their cytokines including IL-4, IL-5, and IL-10.
Dendritic cells (DC) are key drivers of T cell-mediated immune response; vitamin D may have the ability
to inhibit DC maturation. Also, monocytes that are exposed to vitamin D have decreased major
histocompatibility complex (MHC) class II expression and ability to stimulate T cells. Th1 cells are the
primary mediators of graft rejection, while Th2 lymphocytes have a regulatory role
1
.
Inflammatory cytokines are key mediators of GVHD. When patients undergo conditioning for
HSCT, the high intensity chemotherapy and radiation activate host tissues to secrete inflammatory
cytokines, which leads to epithelial cell damage in the gastrointestinal tract. Following this, donor T-cells
are activated, which is characterized by secretion of Th1 cytokines. These cytokines activate mononuclear
phagocytes and ultimately lead to a cytokine storm, which contribute to target cell apoptosis and local
tissue injury
28
. In the setting of compromised gut epithelium during the early post-transplant period,
vitamin D absorption and metabolism are likely to be affected
20
.
Cytokine production can lead to endothelial cell activation and damage throughout the body,
leading to transplant-related complications including veno-occlusive disease (VOD), diffuse alveolar
hemorrhage, transplant-associated thrombotic microangiopathy (TMA), and capillary leak syndrome
3
.
Khandelwal et al conducted a randomized study of the effects of human milk on pro-
inflammatory markers in children undergoing HSCT. Their control group had increased pro-inflammatory
cytokines including IL-6, IL-8, IL-10, and IFN-gamma at day +14
29
.
Caballero-Velazquez et al conducted a prospective study in adult patients undergoing allogeneic
HSCT examining the impact of no vitamin D, low-dose vitamin D (1,000 IU/day), and high-dose vitamin
D (5,000 IU/day) on GVHD, vitamin D levels, immune cells, and cytokines. They found that treatment
with vitamin D significantly decreased the risk of cGVHD. They did not find a significant difference in

22
relapse or survival in control versus treated groups. There also was no significant difference in total
lymphocytes, T cells, or dendritic cells. At day +1, the control group had higher levels of IFN-gamma
than both of the vitamin D groups, and at day +14 the control group had higher levels of IFN-gamma
compared to the high-dose vitamin D group. No significant difference was found in levels of IL-2, IL-4,
IL-6, IL-10, or tumor necrosis factor (TNF)-alpha
19
.
Joseph et al examined vitamin D receptor expression in and the effects of vitamin D on
alloreactive T cells in vitro. They found that alloreactive T cells express high levels of vitamin D receptor,
and that vitamin D can inhibit alloreactivity
30
.
The Institute of Medicine reports 4,000 IU/day as the upper vitamin D supplementation limit for
most individuals, however there is little evidence of toxicity unless doses of 10,000 IU of vitamin D3 per
day or 25-OH vitamin D levels of 150ng/ml are exceeded
31,32
. Multiple studies have raised concern
regarding the adequacy of standard and high dose vitamin D dosing for vitamin D deficiency
33,34
.
Veugelers et al. performed a meta-analysis on vitamin D supplementation and vitamin D status and found
that 2909 IU of vitamin D per day is needed to achieve serum 25-OH vitamin D concentrations of 20
ng/mL or more in 97.5% of healthy individuals. For normal weight, overweight and obese vitamin D
recipients this was 3094, 4450 and 7248 IU respectively. This would translate to about 50 IU/kg/day for
adult individual. There are no comparable data reports in children, especially those with chronic illness
such as HSCT recipients
35
.
Patients with cystic fibrosis (CF) have malabsorption of fat soluble vitamins including vitamin D.
Despite supplementation with high-dose vitamin D (50,000IU/week), patients with CF often still do not
achieve adequate vitamin D levels. Shepherd et al conducted a retrospective chart review comparing
patients with CF who received ultra-high-dose (Stoss, see Table 2.1) vitamin D supplementation followed
by maintenance vitamin D to those who declined Stoss therapy and received supplementation per standard
recommendations (control group). Over a 12-month period, patients who received Stoss therapy had
significantly larger increases in their vitamin D levels compared to the control group (15nmol/L vs
5nmol/L). They found no evidence of vitamin D toxicity
33
.

23
Few studies regarding vitamin D dosing have been done in pediatric patients planned for HSCT.
Wallace et al conducted a study of standard versus high-dose vitamin D supplementation in patients
undergoing HSCT, and found that despite high-dose supplementation (>200IU/kg/day), many patients
failed to achieve therapeutic vitamin D levels
20
. Furthermore, it is challenging in the initial post-
transplant period to reliably administer vitamin D to patients due to their frequently concomitant
mucositis and administration of drugs that interfere with vitamin D absorption. Wallace et al conducted a
prospective pilot study in which Stoss vitamin D supplementation was given to 10 pediatric patients with
vitamin D level <50ng/mL prior to HSCT. All patients achieved a therapeutic vitamin D level of
>30ng/mL. The mean peak serum level of vitamin D was 80.4+28.6ng/mL, which is well below the level
at which toxicity may be seen (150ng/mL). This was well tolerated without any significant side effects
21
.
While Wallace et al found that Stoss dosing can be given safely to patients prior to transplant, to
our knowledge, no studies have been done evaluating the effects of Stoss on biomarkers of inflammation.
We believe this study will have minimal risk, given that Stoss dosing has been given safely in patients
with both CF and those receiving HSCT without any significant side effects
21,33
. Either Stoss dosing or
daily or weekly dosing of Vitamin D is therefore considered routine care for Vitamin D supplementation
prior to HSCT.

Objectives
The primary aims of this study are to (a) evaluate whether Stoss dosing is more effective than
routine supplementation to achieve sufficient pre-HSCT vitamin D levels, and (b) to compare the
incidence of transplant-related complications with a historical cohort, specifically those involving
endothelial injury occurring due to high inflammatory state. The rationale for co-primary aims is that
benefit seen with either aim would be construed as evidence of efficacy (i.e. “success”) for Stoss Vitamin
D supplementation in this population.
21

24
The secondary aim is to evaluate the dynamic changes in inflammatory biomarker (i.e. cytokine
and immunophenotyping) panels during the transplantation process in subjects who receive Stoss therapy
prior to HSCT.

Study methods and procedures
Design
This is an interventional pilot study evaluating the prevalence of pre-HSCT Vitamin D
sufficiency and the incidence of transplant-related events in HSCT recipients with vitamin D level
<50ng/mL who receive single ultra-high dose vitamin D (Stoss) therapy prior to starting the
transplantation process. Enrolled subjects will receive oral Stoss therapy (see Table 2.1) prior to HSCT.
Stoss dosing is one accepted standard method of vitamin D supplementation for various chronic
conditions and in patients receiving HSCT. We will evaluate biomarkers including levels of inflammatory
cytokines at baseline and at regular intervals until day +100, which encompasses the time in which
patients have a high inflammatory state in the early post-transplant period.

Figure 2.1. Study design

25

Figure 2.2. Laboratory monitoring

The study enrollment and specimen collection will take place at CHLA. All research-related
activities will be conducted at sites affiliated with CHLA or USC.
Drug Administration
Once consent and assent (if applicable) are obtained and baseline study samples are collected, a
member of the study team or clinical provider will observe the subject take the prescribed vitamin D dose
during scheduled outpatient or inpatient visit. Vitamin D3 (cholecalciferol) dose will be calculated based
on study subjects’ vitamin D level and weight (kg). Vitamin D3 (cholecalciferol) will be either a liquid
formulation of 5000 units/mL from National Vitamin Company (NDC 54629-0772-41), capsule of 50,000
units/capsule from Nivagen (NDC 75834-0020-01), or capsule of 5,000 units/capsule from Major (NDC
00904-5986-60). It will be administered orally and may be taken with food based on patient’s preference.
To participate in the study, subjects will be required to take the full prescribed dose. It will be dispensed
by a CHLA pharmacist. The drug will be given at no cost to the study subjects. This will be done
approximately 1-2 weeks prior to beginning conditioning for HSCT (exact timing of administration
subject to investigator discretion). This may take place in the outpatient or inpatient setting. See Table
2.1a for dosing guidelines and 1b for potential Vitamin D associated toxicity.

26

Descriptor

Symptom

Frequency
Frequent

---- occurs in 25 or more people
out of 100
Common --- occurs in 10 - 25 people out
of 100
Uncommon Allergy occurs in 1 - 10 people out
of 100
Rare Hypercalcemia occurs in less than 1 person
out of 100
Table 2.1b. Toxicity due to vitamin D

Planned Specimen and Data Collection
Serum 25OHD levels will be obtained as part of routine clinical care approximately weekly, or
with each outpatient visit if less often, until day +100 post-transplant. If patients’ vitamin D levels plateau
and decrease to 30ng/mL, they will be placed on vitamin D supplementation per provider discretion.
Biomarker panel and cytokine stimulation will be obtained as research samples as outlined below.
Samples will be processed and stored until tested in a laboratory affiliated with CHLA or USC. We will
also record results of tests that are standard of care, including serum vitamin D levels, serum calcium,
serum phosphorus, serum PTH, and urine calcium to creatinine. See Table 2.2 for detailed schedule of
assessments and blood volume for research samples.
Research studies
a. Cytokine panel: This is a research sample testing for levels of various cytokines, including but not
limited to IFN-gamma, TNF-alpha, IL-1a, IL-1b, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10. The volume of this is
25-OH D level Single vitamin D3
dose
<10ng/mL 14,000 IU/kg/dose
10-29ng/mL 12,000 IU/kg/dose
30-50ng/mL 7,000 IU/kg/dose
Max total single dose: 600,000 IU
Dose rounded to nearest 5,000 IU
Table 2.1a. Stoss dosing

27
0.2mL of serum per sample, which will require 0.5mL of whole blood. See Table 2.2 for a detailed
schedule of assessments.
b. Immunophenotyping panel: This is an immunophenotyping research sample assessing the frequency of
circulating immune cells and their phospho-protein alterations after stimulation by various cytokines at
hyper-physiologic doses. The volume of whole blood collected per sample is 12mL. See Table 2.2 for a
detailed schedule of assessments.
c. If there are remaining research specimens once the aforementioned tests have been completed, they
may be used for additional testing related to the aims of this study.

28

Screening and
Enrollment
First
baseline
(prior to
Stoss)
Second baseline
(up to 2 weeks
after Stoss, and
before HSCT)
HSCT Day 0 to
Day +100
(weekly)*
Days 0,
+7,+14, and
+30º
Inclusion/ Exclusion Assessment X
Medical history/ Demographics X
Informed Consent X
Serum Vitamin D^ X X X
Serum Ca^ X X X
Serum Phosphorus^ X X X
Serum PTH^ X X X (on day
+30 only)
Urine Ca/Cr^ X X X
Serum Cytokine Panel for
Research
1

X X X
Immunophenotyping Panel
with/without Cytokine
Stimulation for Research
2

X X
AEs/SAEs
3
Per institutional IRB requirements
* Obtained as standard of care approximately every week, or with each outpatient visit if less often, until 100 days.
^These assessments are performed as per institutional standard of care, and results will be collected if available.
Assessment schedule may vary as clinically indicated. Assessments that are not obtained will not constitute a protocol
deviation.
ºAssessments may have a grace period of +/- 3 days. They will be obtained at the times specified unless subjects’
clinical status precludes this.
1
Will obtain up to 3mL of whole blood per sample collected in a red-top tube
2
Will obtain up to 12mL of whole blood per sample collected in a green-top heparinized non-glass tube
3
AEs and SAEs will be monitored until day +100. See section 10 of protocol for further information.
Table 2.2. Schedule of Assessments
Any research samples will be obtained in accordance with institutional standards to ensure patient
safety and without compromising needed diagnostic/clinical blood samples or patient health. The total
amount of blood drawn will not exceed 5% of the estimated blood volume in any 24 hour period. The
total volume of blood in any 24 hour period will include blood drawn for clinical testing, research and
waste samples combined.

29
Blood will be drawn through patients’ existing central lines when available. If patients do not
have a central line that can be accessed, research samples may be obtained via venipuncture (peripheral
needle stick) but this will not be required. Since patients undergoing HSCT have daily laboratory studies
performed as routine clinical care, additional study-related tests will be added on to already collected
blood samples whenever possible. Extra blood would only be drawn if specific collection tubes are
required. Whenever possible, study blood samples will be obtained at the same time as routine blood
draws to eliminate or minimize additional central line access or venipuncture. Urine will be collected by
clean catch or cotton balls placed in the diaper as is routinely done during transplantation so
catheterization will not be required. However, if a catheter is inserted for clinical indications, urine from
the Foley bag will be obtained.
In addition to the aforementioned routine and research samples, we will capture relevant disease
and transplant specific clinical data including but not limited to donor characteristics, transplant regimen,
toxicities, and outcomes.

Outcomes and Endpoints
The primary aims of this study are to (a) evaluate whether Stoss dosing increases pre-HSCT
vitamin D levels better than lower dose daily or weekly supplementation, and (b) observe the incidence of
transplant-related complications, specifically those involving endothelial injury: GVHD (allogenic
HSCT), VOD and TMA (allogenic and autologous HSCT)). The secondary aims are to evaluate (1) the
dynamic changes in inflammatory biomarker panels in subjects who receive Stoss therapy prior to HSCT,
(2) average Vitamin D level and incidence of Vitamin D insufficiency post-HSCT in those who receive
Stoss dosing.
The primary study endpoints are pre-HSCT vitamin D levels ≥30 ng/ml (“sufficient”) and the
incidence of transplant-related events by day +100 after HSCT. For subjects undergoing allogeneic
HSCT, this entails day +100 from first HSCT. For subjects undergoing autologous HSCT, this entails day

30
+100 from primary scheduled transplant therapy. We will observe their clinical status via chart review to
capture any additional significant events until 1 year after HSCT.

Statistical Considerations
Based on retrospective data from our institution, with routine supplementation approximately
66% of patients have sufficient pre-HSCT vitamin D levels (≥30ng/ml). Based on a one-sided test with
type I error of 0.05 with 80% power to detect a significant increase in the prevalence of sufficient pre-
HSCT vitamin D levels from the baseline of 66% to >90%, a sample of 40 subjects is needed.
Based on retrospective data from our institution, the current incidence of GVHD (grade 2-4),
VOD, and TMA in patients here at CHLA is 40%. Based on a one-sided, one-sample test of proportions
with type I error 0.05 with 80% power to detect a significant reduction in the complications rate from the
baseline of 0.40 to a new rate of 0.20, a sample size of 38 subjects is needed.
Due to the short time-frame and need for HSCT, we anticipate no drop-out between enrollment
and evaluation for the pre-HSCT Vitamin D primary aim. Patient dropout prior to Day+100 is most
likely due to toxicity, which constitutes an evaluable event. Due to the longer follow-up, however, the
transplant complication co-primary aim includes an anticipated non-event related dropout rate of <5% (2
patients). These patients will not be replaced on study. We will therefore enroll up to 50 patients in our
study to ensure we have 40 evaluable patients for both primary aims. We will re-evaluate our accrual
goals based on the number of evaluable patients.

Patient Screening, Eligibility, and Enrollment
Study Population
We will identify patients aged 0-25 years of age undergoing evaluation for planned HSCT and
approach them for participation in the study. Vitamin D levels are obtained on all patients prior to HSCT
as our current standard of care. Those patients with 25OHD levels between 0 and 50 ng/mL who have

31
consented will continue on study. We will include subjects with vitamin D level up to and including
50ng/mL because studies have found that a majority of patients who do not have vitamin D deficiency
before transplant develop vitamin D deficiency by day +100
4
. Clinical laboratory studies obtained within
approximately 4 weeks prior to enrollment may be utilized. Stoss dosing has been given in patients with
pre-treatment vitamin D levels up to and including 50ng/mL without significant adverse events
21
.
Inclusion and Exclusion Criteria
Inclusion Criteria: Patients who are preparing for HSCT. If a patient is receiving an autologous
transplant, enrollment must occur prior to first transplant in the case that the patient is planned for
multiple transplants.
Exclusion criteria: Patients with clinically significant uncorrected hypocalcemia or
hypophosphatemia. Patients in the ICU or on renal replacement therapy. Patients who have had an
allogeneic transplant within the past 12 months prior to enrollment.
Consent Process
After consulting with their primary attending, a research team member will approach eligible
subjects and their parent(s)/ legal guardian(s) about the study and will explain the study protocol, risks
and benefits. If the adult subjects and the parent(s)/legal guardian(s) are interested in this study, assent, if
appropriate, and consent will be obtained. There will be no selection by gender. The informed consent
process will be done in the families’ primary language. Recruitment, informed consent, and assent will be
done in a private area.
In subjects < 18 years old, written consent from one parent(s)/legal guardian(s) will be sufficient.
Subjects ages 7 to 17 will be asked to give written assent. Subjects who are ≥ 18 years old will be asked
to give written consent.
Following the subjects’ 18
th
birthday, they will be approached by study personnel at their next
appointment to reconsent them and have their permission to continue per our consent conference standard

32
procedures. Reconsenting must occur before the end of the calendar year. Non-English speaking subjects
will also be enrolled, and study personnel will communicate with them with an authorized interpreter.

Risk/Benefit Assessment
Potential risk to research subjects includes the possibility of overtreatment with cholecalciferol.
Cholecalciferol toxicity is extremely rare as outlined in the background of this protocol. In some cases,
this can lead to increased circulating calcium to higher than normal levels (hypercalcemia), as well as lead
to calcium deposition in the kidneys contributing to nephrocalcinosis or nephrolithiasis. Additionally,
excessive cholecalciferol administration can sequester serum calcium in the bone matrix rapidly resulting
in hypocalcemia (hungry bone syndrome). All subjects will be monitored prior to and frequently during
treatment to identify those at risk for hypervitaminosis D and related effects. Those affected patients will
receive appropriate clinical treatment for these conditions should they occur. Any subject with a urine
calcium/creatinine ratio >0.2 will have further workup to assess for nephrocalcinosis or nephrolithiasis if
clinically indicated (per provider discretion). Since all subjects will receive a HSCT and will be admitted
to the inpatient BMT unit, they will have electrolytes and calcium levels checked nearly daily. We
anticipate that any abnormal calcium levels will be discovered quickly with frequent lab monitoring.
Subjects with evidence of hypercalciuria and/or nephrocalcinosis by ultrasound will be treated per
provider discretion. If hypocalcemia is evident, then either enteral or parenteral calcium will be
administered and if indicated an endocrinology consult will be obtained for any additional management.
There is a potential risk of breech of patient identity and privacy during our study. Study
personnel will employ standard measures to protect patient privacy and the confidentiality of all patient
information.
Participating subjects will benefit from close monitoring and expert interpretation of vitamin D
levels. We feel that the potential benefits to the research participants and future patients justify the

33
exposure of the participants to the risk. As stated above, we believe this intervention carries minimal risk
for participants.
The alternative to participation is not participating, and continuing treatment and monitoring per
current standard of care.

Safety Monitoring
This study is low risk and will not require DSMB monitoring. The CHLA principal investigator
(PI) and PIs designees will monitor the research for the safety of the participants. Timing for monitoring
for serious adverse events (SAE) and adverse events (AE) are outlined in Table 2.2. SAE are defined as
an unexpected event probably or definitely related to vitamin D with the outcome of disability or
permanent damage, hospitalization, life-threatening outcome, congenital anomaly/birth defect, required
intervention to prevent permanent impairment or damage, or other serious important medical events. AE
are defined as events not rising to the level of a SAE. Any unexpected safety events attributed to vitamin
D therapy grade 4 and higher will be reported promptly to the Institutional Review Board (IRB) according
to IRB requirements: SAE will be reported within 5 business days of the responsible member of the study
team discovering the event; SAE regarding death will be reported within 24 hours of the responsible
member of the study team discovering the event. Unexpected safety events probably or definitely
attributed to vitamin D therapy that are grade 3 and lower will be reported at continuing review. Grading
of AE will be done according to CTCAE v4.03. Abnormal laboratory values or test results constitute
adverse events if they induce clinical signs or symptoms or require therapy. However, as a supportive care
study for patient receiving often toxic HSCT regimens, AE and SAE attributed to underlying HSCT
therapy (i.e. not probably or definitely attributed to the Vitamin D itself) will not be routinely reported.
Monitoring and auditing procedures will be followed to ensure that the study is conducted,
documented, and reported in accordance with the IRB approved protocol, all applicable federal
regulations and guidelines, and applicable regulatory requirements of Children’s Hospital Los Angeles.

34
Privacy and Data Confidentiality
Research procedures will be conducted in a private setting. Data will be captured and reviewed in
a private setting. The collection of information about participants is limited to the amount necessary to
achieve aims of the research. Data and/or specimens will be labeled with a code that the research team
can link to personal identifying information. Research data will be kept in a locked office with access
restricted to authorized study personnel. Electronic data will be maintained on a secure computer/laptop
that is password protected and has security software installed. There will be restrictions on copying study
related materials. Access rights will be terminated when authorized study personnel leave the study team.
Coded data and/or specimens will not be released to a third party. Research data and/or specimens will be
retained for study record keeping purposes per institutional policy.

Financial Obligations and Funding
There will be no direct cost to study participants for participation in this study. Study participants
will not be charged for high dose Vitamin D. The laboratory studies obtained in the course of this study
are standard of care for clinical monitoring of vitamin D deficiency during the HSCT process and will be
covered by the subjects insurance with the exception of research samples used for biomarker analysis,
which will be performed at no cost to the study participant. Subjects or their insurance companies will be
charged for hospitalizations including tests and treatments of any side effects of high dose vitamin D. In
the event that subjects’ insurance does not cover these costs, subjects may incur additional costs because
of treatment side effects.
Participants will not be reimbursed for their participation in this study.
This study will be funded by departmental funds and a CHLA Core Pilot Award.

35
Status of Clinical Trial to Date
Enrollment
This study received IRB approval from CHLA in October 2018. Enrollment began in December
2018 and completed in January 2020. Thirty-three subjects have been enrolled. This accrual was delayed
relative to our goal (Figure 2.3).

Figure 2.3. Study accrual as of January 2020

Preliminary Results
Of the 33 enrolled subjects, three were withdrawn (two were unable to complete necessary study
activities prior to HSCT, and one died before beginning study activities) and one subject’s HSCT was
delayed until further notice. The remaining 29 subjects had a significant increase in mean vitamin D level
following Stoss dosing, from a baseline pre-Stoss vitamin D level of 27.7 to post-Stoss vitamin D level of
70.2 (p<0.001, paired samples t-test). Evaluation of the association between vitamin D level, cytokines,
immunophenotyping panels, and post-HSCT complications is ongoing.
Adverse Events
There have been no SAEs attributable to vitamin D. The peak vitamin D level was 112ng/mL,
which is below the level concerning for toxicity. There were no incidences of nephrolithiasis or
0
5
10
15
20
25
30
35
40
45
Dec-18
Jan-19
Feb-19
Mar-19
Apr-19
May-19
Jun-19
Jul-19
Aug-19
Sep-19
Oct-19
Nov-19
Dec-19
Jan-20
Number of Subjects Enrolled
Study Accrual
Goal
Actual

36
hepatopathy attributable to vitamin D supplementation. One subject vomited the dose shortly after taking
it (grade 1 toxicity). Otherwise, the dose was well tolerated.
Barriers to Enrollment
The primary barrier to enrollment has been the timing of research-related activities prior to
HSCT. Potential subjects must be approached at least one week prior to admission for HSCT, and must
have subsequent planned visits to complete the remainder of the research activities in a timely manner.
The vitamin D level is currently a send out test that takes up to five days to result. As CHLA is a large
referral center, some patients have their pre-HSCT evaluations done at other facilities and have very few
appointments at CHLA prior to HSCT.
The successes and barriers of this study have provided valuable insight and informed our
approach for future clinical trials. In future trials, we will strive to incorporate routine clinical studies (e.g.
vitamin D level) with shortened turnaround time. We will also consolidate research-related activities to
decrease the amount of visits that are needed to complete the activities prior to HSCT.

Conclusions
The study presented in Chapter 1 noted that in a retrospective study of 314 pediatric patients, we
found an association between vitamin D levels and overall outcomes following HSCT. We also found that
low-dose vitamin D supplementation does not adequately achieve or maintain vitamin D sufficiency,
necessitating revisiting both our vitamin D supplementation and assessment practices. The HSCT patient
population is particularly vulnerable to vitamin D insufficiency and is refractory to low-dose
supplementation, therefore, an alternative dosing regimen such as Stoss is beneficial.
Preliminary study results in Chapter 2 show that Stoss dosing effectively achieves vitamin D
sufficiency in pediatric patients prior to HSCT, without any SAEs attributable to vitamin D. We anticipate
that the final results of our study will enhance our understanding of the immunomodulatory effects of
vitamin D, specifically in the context of endothelial injury. Further investigation is needed to better

37
elucidate the relationship between vitamin D levels and post-HSCT complications and outcomes. This
trial serves as a foundation for larger prospective studies to examine the interaction between the immune
response and post-HSCT complications, as well as strategies to mitigate them.

38
References
1. van Etten E, Mathieu C. Immunoregulation by 1,25-dihydroxyvitamin D3: basic concepts. J
Steroid Biochem Mol Biol. 2005;97(1-2):93-101.
2. Arain A, Matthiesen C. Vitamin D deficiency and graft-versus-host disease in hematopoietic stem
cell transplant population. Hematol Oncol Stem Cell Ther. 2018.
3. Carreras E, Diaz-Ricart M. The role of the endothelium in the short-term complications of
hematopoietic SCT. Bone marrow transplantation. 2011;46(12):1495-1502.
4. Wallace G, Jodele S, Howell J, et al. Vitamin D Deficiency and Survival in Children after
Hematopoietic Stem Cell Transplant. Biology of blood and marrow transplantation : journal of
the American Society for Blood and Marrow Transplantation. 2015;21(9):1627-1631.
5. Beebe K, Magee K, McNulty A, et al. Vitamin D deficiency and outcomes in pediatric
hematopoietic stem cell transplantation. Pediatric blood & cancer. 2018;65(2).
6. Hansson ME, Norlin AC, Omazic B, et al. Vitamin d levels affect outcome in pediatric
hematopoietic stem cell transplantation. Biology of blood and marrow transplantation : journal of
the American Society for Blood and Marrow Transplantation. 2014;20(10):1537-1543.
7. Macris P. Hematopoietic Stem Cell Transplantation Nutrition Care Criteria. 3rd ed. Seattle:
Seattle Cancer Care Alliance; 2012.
8. Bischoff-Ferrari HA. Optimal serum 25-hydroxyvitamin D levels for multiple health outcomes.
Adv Exp Med Biol. 2008;624:55-71.
9. Khan QJ, Fabian CJ. How I treat vitamin d deficiency. J Oncol Pract. 2010;6(2):97-101.
10. Holick MF, Binkley NC, Bischoff-Ferrari HA, et al. Evaluation, treatment, and prevention of
vitamin D deficiency: an Endocrine Society clinical practice guideline. J Clin Endocrinol Metab.
2011;96(7):1911-1930.

39
11. Atkinson K, Horowitz MM, Gale RP, Lee MB, Rimm AA, Bortin MM. Consensus among bone
marrow transplanters for diagnosis, grading and treatment of chronic graft-versus-host disease.
Committee of the International Bone Marrow Transplant Registry. Bone marrow transplantation.
1989;4(3):247-254.
12. Przepiorka D, Weisdorf D, Martin P, et al. 1994 Consensus Conference on Acute GVHD
Grading. Bone marrow transplantation. 1995;15(6):825-828.
13. Jones RJ, Lee KS, Beschorner WE, et al. Venoocclusive disease of the liver following bone
marrow transplantation. Transplantation. 1987;44(6):778-783.
14. McDonald GB, Hinds MS, Fisher LD, et al. Veno-occlusive disease of the liver and multiorgan
failure after bone marrow transplantation: a cohort study of 355 patients. Ann Intern Med.
1993;118(4):255-267.
15. Jodele S, Davies SM, Lane A, et al. Diagnostic and risk criteria for HSCT-associated thrombotic
microangiopathy: a study in children and young adults. Blood. 2014;124(4):645-653.
16. Stata Statistical Software [computer program]. College Station, TX: StataCorp LP; 2009.
17. Copelan E, Casper JT, Carter SL, et al. A scheme for defining cause of death and its application
in the T cell depletion trial. Biology of blood and marrow transplantation : journal of the
American Society for Blood and Marrow Transplantation. 2007;13(12):1469-1476.
18. Sproat L, Bolwell B, Rybicki L, et al. Vitamin D level after allogeneic hematopoietic stem cell
transplant. Biology of blood and marrow transplantation : journal of the American Society for
Blood and Marrow Transplantation. 2011;17(7):1079-1083.
19. Caballero-Velazquez T, Montero I, Sanchez-Guijo F, et al. Immunomodulatory Effect of Vitamin
D after Allogeneic Stem Cell Transplantation: Results of a Prospective Multicenter Clinical Trial.
Clinical cancer research : an official journal of the American Association for Cancer Research.
2016;22(23):5673-5681.

40
20. Wallace G, Jodele S, Myers KC, et al. Vitamin D Deficiency in Pediatric Hematopoietic Stem
Cell Transplantation Patients Despite Both Standard and Aggressive Supplementation. Biology of
blood and marrow transplantation : journal of the American Society for Blood and Marrow
Transplantation. 2016;22(7):1271-1274.
21. Wallace G, Jodele S, Myers KC, et al. Single Ultra-High-Dose Cholecalciferol to Prevent
Vitamin D Deficiency in Pediatric Hematopoietic Stem Cell Transplantation. Biology of blood
and marrow transplantation : journal of the American Society for Blood and Marrow
Transplantation. 2018.
22. Zhang Y, Leung DY, Richers BN, et al. Vitamin D inhibits monocyte/macrophage
proinflammatory cytokine production by targeting MAPK phosphatase-1. J Immunol.
2012;188(5):2127-2135.
23. McNally JD, Nama N, O'Hearn K, et al. Vitamin D deficiency in critically ill children: a
systematic review and meta-analysis. Crit Care. 2017;21(1):287.
24. von Bahr L, Blennow O, Alm J, et al. Increased incidence of chronic GvHD and CMV disease in
patients with vitamin D deficiency before allogeneic stem cell transplantation. Bone marrow
transplantation. 2015;50(9):1217-1223.
25. Glotzbecker B, Ho VT, Aldridge J, et al. Low levels of 25-hydroxyvitamin D before allogeneic
hematopoietic SCT correlate with the development of chronic GVHD. Bone marrow
transplantation. 2013;48(4):593-597.
26. Campos DJ, Boguszewski CL, Funke VA, et al. Bone mineral density, vitamin D, and nutritional
status of children submitted to hematopoietic stem cell transplantation. Nutrition. 2014;30(6):654-
659.
27. Bhandari R, Malvar J, Sacapano A, Aguayo-Hiraldo P, Jodele S, Orgel E. Association between
Vitamin D and Risk for Early and Late Post-Transplant Complications. Biology of blood and
marrow transplantation : journal of the American Society for Blood and Marrow
Transplantation. 2020;26(2):343-350.

41
28. Ferrara JL, Reddy P. Pathophysiology of graft-versus-host disease. Semin Hematol. 2006;43(1):3-
10.
29. Khandelwal P, Andersen H, Taggart C, et al. A Randomized Trial of Human Milk to Maintain
Microbiome Diversity and Reduce Intestinal Inflammation during Stem Cell Transplantation.
Biology of blood and marrow transplantation : journal of the American Society for Blood and
Marrow Transplantation. 2018;24(3):S85-S86.
30. Joseph RW, Bayraktar UD, Kim TK, et al. Vitamin D receptor upregulation in alloreactive human
T cells. Human immunology. 2012;73(7):693-698.
31. Ross AC, Manson JE, Abrams SA, et al. The 2011 report on dietary reference intakes for calcium
and vitamin D from the Institute of Medicine: what clinicians need to know. J Clin Endocrinol
Metab. 2011;96(1):53-58.
32. Hathcock JN, Shao A, Vieth R, Heaney R. Risk assessment for vitamin D. Am J Clin Nutr.
2007;85(1):6-18.
33. Shepherd D, Belessis Y, Katz T, Morton J, Field P, Jaffe A. Single high-dose oral vitamin D3
(stoss) therapy--a solution to vitamin D deficiency in children with cystic fibrosis? J Cyst Fibros.
2013;12(2):177-182.
34. Rajakumar K, Moore CG, Yabes J, et al. Effect of Vitamin D3 Supplementation in Black and in
White Children: A Randomized, Placebo-Controlled Trial. J Clin Endocrinol Metab.
2015;100(8):3183-3192.
35. Veugelers PJ, Pham TM, Ekwaru JP. Optimal Vitamin D Supplementation Doses that Minimize
the Risk for Both Low and High Serum 25-Hydroxyvitamin D Concentrations in the General
Population. Nutrients. 2015;7(12):10189-10208.

Abstract (if available)

Abstract Hematopoietic stem cell transplantation (HSCT) is a therapeutic and often curative approach to pediatric blood disorders and cancers. While HSCT outcomes continue to improve by means of reduced intensity conditioning regimens and alternative donor sources, complications such as graft-versus-host disease (GVHD) and veno-occlusive disease (VOD) are often severe and at times fatal. These complications are mediated by inflammation and endothelial injury. The vitamin D (VD) receptor is present on all cells. VD has immunomodulatory effects, and may play a role in mitigating the inflammatory response that is implicated in processes such as GVHD and VOD. While the relationship between VD and post-HSCT complications requires further investigation, studies have found an association between VD deficiency and GVHD. Additionally, multiple studies have found that prior to HSCT, a majority of pediatric patients have VD insufficiency that is not improved with low-dose supplementation. We investigated the association between VD and post-HSCT complications, also evaluating practices of VD monitoring and supplementation (Chapter 1). This was accomplished through a retrospective study of pediatric patients who received HSCT between January 2013 and April 2018. We found that the majority of patients were VD insufficient prior to HSCT, and low-dose supplementation did not significantly increase VD levels. We did not find an association between VD and GVHD or VOD, but there was a correlation between VD and overall survival. Building on these findings, we are prospectively investigating an alternative VD dosing regimen, further characterizing the relationship between VD and the inflammatory response, and evaluating the relationship between VD and post-HSCT complications. In this ongoing study (Chapter 2), subjects receive a single oral ultra-high-dose of VD (Stoss) prior to HSCT. Immunophenotyping panels and cytokine levels are obtained prior to and up to 30 days following Stoss dosing to characterize the change in immune response with Stoss dosing. VD levels and incidence of GVHD and VOD are monitored for 100 days following Stoss dosing. Preliminary results show that Stoss dosing significantly increases VD levels prior to HSCT. These two studies help us to better understand the immunomodulatory role of VD in HSCT and the efficacy of a regimen such as Stoss dosing in achieving VD sufficiency in pediatric patients receiving HSCT. Further investigation is needed to determine the effect of VD on post-HSCT complications.

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Asset Metadata

Creator Bhandari, Rusha Hiren (author)

Core Title The role of vitamin D in pediatric hematopoietic stem cell transplantation

School Keck School of Medicine

Degree Master of Science

Degree Program Clinical, Biomedical and Translational Investigations

Publication Date 05/07/2020

Defense Date 05/05/2020

Publisher University of Southern California (original), University of Southern California. Libraries (digital)

Tag hematopoietic stem cell transplantation,OAI-PMH Harvest,pediatric,vitamin D

Language English

Contributor Electronically uploaded by the author (provenance)

Advisor Orgel, Etan (committee chair), Asgharzadeh, Shahab (committee member), Pulsipher, Michael (committee member)

Creator Email rbhandari@chla.usc.edu,rushabhandari1@gmail.com

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