Download The role of temperature in cancer immunology

Chronic Mild Thermal Stress Stimulates Tumor
Growth and Depresses the Anti-Tumor Immune
Response
Elizabeth Repasky, Ph.D.
Professor of Immunology,
Program Leader- Cell Stress and Biophysical Therapies Program
Roswell Park Cancer Institute- Buffalo, NY USA
[email protected]
ICNIRP/WHO Workshop
“A Closer look at the thresholds of thermal damage”
Istanbul, Turkey, May 26-28, 2015
How ıs the ımmune system
regulated by body temperature?
Mild (Fever-Range) Hyperthermia Affects Many
Aspects of the Tumor Microenvironment
Repasky, Evans, Dewhirst,
Cancer Immunology Research, 2013
3
THERMOREGULATION- A unique automomic
system
• Evolved to regulate a stable core over wide
range of ambient temperatures, heat loads
from work, and fever
• One of the only systems that also relies on
behavior and conscious awareness of the
environment to achieve regulation
Effectiveness of HumanThermoregulation
Downloaded from: StudentConsult (on 13 May 2007 10:19 AM)
© 2005 Elsevier
The stress
response
Outline
• Integrated definition
– Stressor (internal or external stimulus)
– Stress perception in the brain
– Physiologic adaptions ("fight-or-flight")
• Internal stimuli
– Emotions (depression, fear)
– Anxiety (crowding, isolation)
Sympathetic Nervous
System (SNS)Norepinephrine
(adrenalin)
Hypothalamic-PituitaryAdrenal axis (HPA-axis)
• External stimuli
– Chemical (toxins)
– Biological (infection, inflammation, metabolic)
– Physical (temperature, pain)
Dhabhar et al, Immunol Res 2014
Joel et al, Nature Reviews Neuroscience 2009
Glaser et al, Nature Reviews Immunology 2005
CRV= corticotropin-releasing hormone
AVP= arginine vasopressin
STRESSORS
Hormonal and
“hard-wired”
mediators of
stress
CNS Cytokines
Hypothalamus
CRV
AVP
ACTH
Adrenal
Glands
Vagus
nerve
SNS
Glucocorticoids
Immune
Cells
Thymus
Spleen
Bone
Marrow
Paul- Fundamental Immunology
Norepinephrin
(NE) and
B-adrenergic
receptors
Lymph
Nodes
Immune System
(Organs & Cells
PNS
Sensitivity to ambient temperature
depends on surface/volume ratio-
~37 oC
Mice are very sensitive to ambient temperature
~37 oC
~37 oC
Surface Area to
Volume Ratio
6
3
1.5
Mice easily gain, or lose, heat from their
environment
Adapted from Blumberg, Body Heat: Temperature
and Life on Earth, 2002
Housing temperature for laboratory mice—
An unexpected tool for studying cold stress and cancer
• The thermoneutral temperature for laboratory mice is
between 29-31°C (~84-88°F)
• At thermoneutrality, little or no extra metabolism is
required to maintain body temperature.
(Gordon, Thermoregulation in Rodents, 1993; Cannon, Physiological Revs., 2004)
• But IACUC certified facilities must select and maintain
a temperature between 20-26°C (~68-78°; until 2011,
18-24 C permitted). (Guide for the Care and Use of Laboratory Animals,
8th Ed, 2011)
• The standard temperature ranges in Europe and Asia are
even lower (approx.18-24°C).
• The temperature in the animal facility at RPCI is maintained
between 22-23°C (approx. 72 oC)
Mice in a huddle- A typical scene in the
mouse room at ST
Photo by K. Kokolus and S. Sexton,
RPCI
Body Temperature (°C)
Mice at ST and TT maintain a normal body
temperature
n=3 per group
Kokolus et al, PNAS 2013
STATIC DISCHARGE RATE OF CUTANEOUS
WARM AND COLD FIBERS
●
●
30 ºC
20 ºC
Pain threshold
Hensel, 1969
Metabolic Rate
(O2 consumption)
Maintaining a Normal Body Temperature (37oC) in Mice
Requires a Significant Increase in Metabolic Rate when
Housed ST
Basal Metabolic Rate
Standard Room Temperature (22oC)
Thermoneutral Temperature (30oC)
0
10
20
30
Ambient Temperature (°C)
Adapted from Specter et al, Handbook of Biological Data 1956
Silva et al, Physiological Reviews 2006
Canon et al, Journal of Experimental Biology 2011
Cold Stress Induces Norepinephrine Production
to Facilitate Heat Production
∆HEAT
↑UCP1
The uncoupling protein in
mitochondria drives
adaptive thermogenesis
Cold Stress
Nguyen et al, Nature 2011
Macrophages
Cannon et al, Physiology Reviews 2006
Adapted from Karp et al, Journal of Experimental Medicine 2012
4T1BALB/c
Tumor-Free
Tumor-Bearing
80 80
1 hour
28º
38º
% of Time Spent at Temperature
% of Time Spent at Temperature
Laboratory
feel even
more cold after
Usingmice
a thermal
preference
implantation!
apparatustumor
to assess
thermal comfort
34º
*
60 60
22º
*
28º
40 40
20 20
30º
38º
22º
34º
0 0
22 22
28 28
30 30
34 34
38 38
Ambient
Temperature
(°C)
Ambient
Temperature
(°C)
* p>0.001
* p<0.001
30º
Katie Kokolus, PhD
n=5, p=0.01
Older mice (> 18 months) prefer a warmer
temperature (~34 oC)
Bonnie Hylander, PhD
Plasma and Tumor Norepinephrine Levels are
Increased in Mice Housed at ST compared with TT
Naïve and tumor-bearing
mice maintained at ST and
TT for 3-4 weeks
Collect plasma
and tumor lysates
Analyze by ELISA
for norepinephrine
concentrations
C57BL/6 (Naïve):Plasma
pg of NE/mL of serum
20
15
*
10
n=13
5
n=6
*p<0.05
**p<0.01
0
n=5
n=6
p=0.19
p=0.17
18
n=6
n=5
How is tumor growth influenced by ambient
temperature?
Standard Temperature (ST)  22°C (~72°F)
Thermoneutral Temperature (TT)  30°C (~86°F)
• Ambient temperature consistently maintained
• Humidity controlled with an air pump
• 12 hour light/dark cycle (6 AM – 6 PM)
Tumor
injection
ST
ST
TT
TT
2 week acclimation
Monitor tumor
growth and
investigate
immunological
endpoints
Alleviating Cold Stress by Housing Mice at
TT Significantly Improves Tumor Control
Tumor Volume (mm3)
600
4T1
(BALB/c)
B16.F10
(C57BL/6)
Pan02
(C57BL/6)
400
**
***
**
200
0
0
10
20
30
40
n=5-6/group
Kokolus et al, PNAS 2013
*p<0.05
** p<0.01
*** p<0.001
**** p<0.0001
20
Tumor growth is significantly slower in mice housed at
thermoneutrality and this depends on the immune
system
4T1
(Balb/C)
4T1
(SCID)
* p < 0.05
**** p < 0.0001
Kokolus et al, PNAS 2013
21
Mice Housed at TT Develop Fewer Metastastic
Tumors (4T1 tumor model)
*
30
Metastatic Area (% )
Lung W eight (g)
1.5
1.0
0.5
0.0
20
10
0
ST
TT
Standard Temperature
Kokolus et al., PNAS 2013
*
ST
TT
Thermoneutral Temperature
Carcinogen (methylcholanthrene) induced tumor
formation is prevented or delayed at TT
50 µg
MCA
Monitor
tumor
incidence
% Tumor-Free
BALB/c
N = 5; # p < 0.05
CD8+ T Cells in the Tumor Microenvironment are
Increased at TT
ST
TT
20
# Cells/Field (40X)
***
15
10
5
0
CD8
N = 5 - 6; * p < 0.05, *** p < 0.001
Kokolus et al.,- PNAS 2013
Anti-tumor
immunity
Immune
Suppression
Are immunosuppressive cells impacted by
cold stress?
25
Myeloid derived suppressor cells suppress
antitumor immunity through a variety of diverse
mechanisms.
Ostrand-Rosenberg and Sinha 2009
Decreased Spleen Mass in Tumor Bearing Mice at TT
Mass of Spleen (g)
0.5
0.4
0.3
0.2
0.1
0.0
**
ST
TT
Flow Cytometry (Spleen)
Absolute # CD11b+ GR1+ Cells
The spleen is a critical repository for MDSCs
prior to their trafficking to the tumor
microenvironment. (Cortez-Retamozo et al.,
2012)
Significant Reduction in
MDSC in Tumor Bearing
Mice at TT
4.010 5
**
3.010 5
2.010 5
1.010 5
0
CT26
N=10/group
** p<0.01
Kokolus et al., PNAS 2013
Does blocking β-adrenergic signaling in
mice at ST or TT impact tumor growth?
Propranolol
(β-blocker)
Propranolol-treatment converts the ST
phenotype to that seen at TT
Effect depends upon presence of immune system!
(A)
(B)
BALB/c - 4T1 Tumor
Mark Bucsek, Katie Kokolus
SCID - 4T1 Tumor
The sympathetic system can impact immune
Outline
cells via mechanisms
now being defined
(internal)
SN
S
(external)
HPAaxis
Adapted from Bellinger DL and Lorton D, Autonom Neuro 2014
Conclusions: Ambient temperature
impairs the anti-tumor immune response
– Chronic thermal stress results in reduced
CD8+ T cell infiltration and increased
Treg/MDSC
– Increased levels of NE in tumors
– Can be reversed by TT or β-blockers
– Causes environmentally induced resistance
to cytotoxic therapies
• Thermal damage may include the impact of
chronic mild thermal stress
Growing recognition of an ominous role for
norepinephrine in tumor progression
•
Adrenergic signaling protects ovarian tumor cells from anoikisapoptosis due to detachment (Sood et al 2010; Cole & Sood, Clin Canc Res,
2012)
• NE increases VEGF, IL-6, and MMPs in ovarian and pancreatic
cancers leading to increase metastasis. (Thakar et al, Nature Medicine
2006; Lutgendorf et al, Clinical Cancer Research 2003; Guo et al, Oncology Reports
2009)
• NE downregulates MHC class 1 and B7-1 on tumor cells leading to
immune escape. (Wang et al, Plos One 2009)
• NE enhances prostate tumor survival by upregulating the
expression of MAPK, and inactivating the apoptotic molecule,
BAD. (Sastry, JBC 2007; Hassan et al, JCI 2012)
• Activation of β-adrenergic receptors promotes the formation of
primary tumors and metastases. (Campbell et al PloS One 2012; Magnon
et al Science 2013)
Thanks to:
Katie Kokolus, PhD (Postdoc with Todd Schell at Penn State Hershey)
Jason Eng, PhD (MD/PHD Student, now back in Medical School,
SUNY Buffalo
Maeghan Capitano, PhD (Postdoc with Hal Broxmeyer, Indiana Univ.)
Chen-Ting (Kelly) Lee, PhD (Postdoc with Mark Dewhirst, Duke Univ.)
Mark Bucsek (MD/PhD student)
Bonnie Hylander, PhD
Scott Abrams, PhD
Sandra Sexton, VDM
Kelvin Lee, MD/PhD
Chris Gordon, PhD (EPA, NC)
Roswell Park Alliance
Foundation
Rowley NYS Breast
Cancer Program
Lloyd Foundation for
Melanoma Research