Collaborating to combat cancer
Could you provide some background
as to how you came to work together?
Has collaboration accelerated the
advancement of your research?
MH: Dr Szabo wanted to work on the role of
H2S in cancer and found our group on campus. I
already knew of his work on H2S but had no idea
he wanted to expand into cancer biology. I feel
that by uniting our expertise, we subsequently
truly reached a new and exciting phase in
both of our careers. As a direct result of the
synergy of innovation that our collaboration
has created, we have initiated several new lines
of experimental and clinical investigations in
the past year. We are also working very hard to
obtain grant funding for this work, both on the
level of basic science and the level of translating
our findings into potential future anticancer
therapies. Collaboration has greatly enhanced
our competitiveness in this effort.
Dr Szabo, you initially discovered new
functions for H2S in blood vessel regulation
and energy production at the Department of
Anaesthesiology, University of Texas Medical
School. How have you translated this
finding for use in the field of oncology?
and out of cells and travel significant distances
in the body.
CS: When we started these studies, one of
the initial hypotheses we wanted to explore
was a little different (and, ultimately, probably
incorrect). We were wondering if the tumours,
which are known to produce vascular endothelial
cell growth factor (VEGF), will, in turn, stimulate
the surrounding blood vessels to produce H2S,
and thereby induce new blood vessel formation
around the tumour. Instead we found this
unexpected, marked ability of the tumour itself
to make large amounts of H2S, which has the
same function as growing blood vessels around
the tumour – but it does this directly, without the
need for any intermediary factors.
You detected that colorectal cancer
cells contain high levels of the H₂Sproducing enzyme cystathionine-betasynthase (CBS). By what means was this
discovery unearthed?
Dr Hellmich, as a seasoned expert
in cancer cell growth, cancer
microenvironment and metastasis, what
has been your role in elucidating the
influence of H2S on cancer development?
MH: Prior to starting our collaborative studies,
I worked on the interaction of the cancer cell
and its environment in various contexts. I was
always fascinated by the multiple layers of
communication between the tumour cell and
its environment (endothelial cells, fibroblasts
and other cell types). Ultimately this interaction
is what is so crucial for tumour growth and
metastasis. I have conducted work in discovering
new roles of a certain class of chemokine
mediators and in the area of tumour stem cell
biology, which is another key aspect on how
tumours grow and induce metastasis. Thus,
based on my prior work, I was not at all surprised
when Csaba started to talk about the possible
role of H2S in the tumour microenvironment.
I immediately recognised the potential
importance of this mechanism, given the fact
that H2S is essentially a ‘local hormone’, with the
advantage of being a gas, and able to diffuse in
MH: Myself and my surgical oncologist
collaborator, Dr Celia Chao, had a collection
of cancer tissues, and corresponding normal
tissues in their freezer. These materials were
collected from patients undergoing surgery in
our University. Under our joint guidance, Dr
Ciro Coletta, one of the postdoctoral fellows
in Szabo’s laboratory, started to analyse these
tissues for H2S production and for the levels of
the various H2S-producing enzymes. Only one
of three H2S-producing enzymes, CBS showed a
marked increase in colorectal tumour tissue; the
other enzymes were unchanged when compared
to the non-cancerous colon tissue.
DRS CSABA SZABO & MARK HELLMICH
In the context of their successful partnership, Drs Csaba Szabo and Mark Hellmich
highlight how scientists from different research backgrounds can come together to
accelerate scientific progress and develop novel medical therapies in the fight against cancer
You are currently working to develop
methods to block cancer cell pathogenesis
with a view to translating this research for
clinical use. Could you discuss some of the
techniques being explored?
CS: We have expressed the human CBS protein
in vitro and we are currently screening chemical
libraries for potential inhibitors. There are
also known ‘lead’ molecules, which we plan
on optimising for cancer therapy. This is yet
another direction of study we are working on
right now. Once we identify a compound that is
an effective inhibitor of CBS, with low toxicity,
we will test its anti-tumour potential in mice
bearing tumour tissue from humans. Some of
this work will be conducted in the frame of a
new spin-off company we have established, CBS
Therapeutics. We are currently in discussions
with potential investors to support this work.
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DRS CSABA SZABO & MARK HELLMICH
The smell of success
A collaborative approach to understanding cancer has led
researchers at the University of Texas Medical Branch, Galveston, to
uncover a role for the characteristically pungent gas, hydrogen sulphide in
the provision of energy to support cancer growth, cell proliferation and invasion
HYDROGEN SULPHIDE (H2S) is a colourless,
flammable and hazardous gas with a distinct
‘rotten egg’ odour. It occurs naturally in
petroleum, natural gas and hot springs and
is released in large quantities during volcanic
eruptions. Furthermore, it is often produced from
the anaerobic bacterial breakdown of organic
matter, leading to its more common names:
‘sewer gas’ or ‘swamp gas’. Perhaps surprisingly,
considering its toxic nature, H2S is also produced
in the human body in small quantities by most
cells. Three enzymes are known to synthesise H2S:
cystathionine-gamma-lyase (CSE), cystathioninebeta synthase (CBS) and 3-mercaptopyruvate
sulphurtransferase (3-MST). It is synthesised
from the substrate L-cysteine and, because of its
gaseous nature, diffuses to its signalling target
within the source cell or across cell membranes
into nearby cells. By entering the bloodstream,
H2S can also affect remote sites in the body. Its
signalling functions – via various mechanisms
including the activation of potassium membrane
26INTERNATIONAL INNOVATION
channels, stimulation of kinase pathways and
the inhibition of phosphodiesterase enzymes –
induce blood vessel relaxation and angiogenesis;
regulation of neuron communication in the brain;
and stimulation of mitochondria – the energy
producing organelle of the cell.
Dr Csaba Szabo, Professor of Anaesthesiology,
has been studying the role of H2S in critical
illness with his group at the University of Texas
Medical Branch (UTMB), Galveston. A recent
collaboration with colleague Dr Mark Hellmich,
Professor of Surgery, Physiology and Biophysics,
has uncovered that H2S plays a key role in colon
cancer metabolism. Their work, published last
year in Proceedings of the National Academy
of Science (PNAS), entitled ‘Tumour-derived
hydrogen sulphide, produced by cystathioninebeta-synthase, stimulates bioenergetics, cell
proliferation and angiogenesis in colon cancer’,
raises the potential for developing novel therapies
to target this serious disease.
HYDROGEN SULPHIDE:
THE FACTS
FORMULA: H2S
MOLAR MASS: 34.0809 G/MOL
DENSITY: 1.36 KG/M³
BOILING POINT: -60 °C
MELTING POINT: -82 °C
SOLUBLE IN: WATER
CLASSIFICATION: SULPHIDE,
SULPHUR COMPOUNDS
CHANGING DIRECTION
The two researchers began their investigations
to test the hypothesis that tumour cells were
acting via vascular endothelial cell growth
factor (VEGF) to stimulate the surrounding
blood vessels to produce H2S and thus induce
the growth of new blood vessels to meet the
needs of the growing tumour. In fact, what
they found was that one of the H2S-producing
enzymes, CBS, was markedly elevated in colon
cancer cells in comparison to the surrounding,
non-cancerous tissue; resulting in increased H2S
production. This was demonstrated in Szabo’s
laboratory by Dr Ciro Coletta, a postdoctoral
fellow, who was asked by Szabo and Hellmich
to compare the levels of H2S-producing
enzymes and H2S in tissue collected from
cancer patient biopsies and patient-matched
healthy tissue. His experiments showed that,
of the three enzymes, CBS was the only one
that was elevated above background levels.
INTELLIGENCE
Following this significant finding, further in
vitro experiments demonstrated that cultured
colon cancer-derived epithelial cell lines also
exhibited up-regulation of CBS and increased
H2S production compared to non-malignant
colon cells.
Inhibiting CBS activity
almost completely
blocked cancer growth by
eliminating its fuel supply
HYDROGEN SULPHIDE, A TUMOUR
CELL SURVIVAL FACTOR AND FUTURE
ANTICANCER TARGET
OBJECTIVES
To develop new approaches to block H2Ssupported growth and proliferation in the design
of new anticancer therapeutic approaches.
MECHANISMS OF ACTION
FUNDING
In order to understand why the cancer cells are
producing these significant amounts of H2S and
uncover the role of the signalling molecule, the
collaborating labs carried out a series of in vitro
and in vivo studies to determine its localisation,
associating cellular components and functions.
Through separating and isolating particular
types of cell component and organelle – using
a process known as cell fractionation – the
researchers were able to demonstrate that
a large proportion of the total cellular CBS
was associated with the mitochondria in
colon cancer cells, specifically to the outer
mitochondrial membrane. The significance of
this finding was supported by evidence that
H2S can stimulate mitochondrial function to
increase the cellular energy production that is
necessary for tumour cell growth, replication
and survival.
Institute for Translational Sciences at the
University of Texas Medical Branch, supported
in part by a Clinical and Translational Science
Award (UL1TR000071) from the National
Center for Advancing Translational Sciences
(NCATS), National Institutes of Health (NIH)
Additional in vitro experiments carried out by
the researchers highlight the importance of
H2S for tumour cell proliferation, migration
and invasion. By using short-hairpin RNA
sequences to suppress the expression of CBS
in colon cancer-derived cell lines, and thus
inhibit production of H2S, the researchers
significantly reduced cell proliferation and
growth of tumours; an effect that was not
replicated in cells from non-cancerous tissue.
Conversely, engineered over-production of CBS
resulted in an increase in the rate of cancer cell
proliferation and invasion.
The accumulated evidence collected by the
UTMB researchers has led them to conclude
that the cancer cell-produced H2S serves three
functions: “First, it stimulates cell division,
perhaps by increasing the activity of specific
intracellular signalling pathways. Second, it
serves as a fuel to feed cancer cell metabolism
by providing the mitochondria with electrons
necessary for energy production. Finally, H2S
was found to diffuse outside the tumour
cells to promote the growth of new blood
vessels to supply the growing tumour tissue,”
explains Szabo.
FROM BENCH TO BEDSIDE
The results from the colon cancer studies
suggest that CBS could represent a therapeutic
target for anticancer drugs. Experiments carried
out by researchers from Hellmich’s laboratory
have demonstrated that pharmacologically
inhibiting CBS activity almost completely
blocked cancer growth by eliminating its fuel
supply. The potential of this is clear: “The
goal would be to block H2S in the tumours,
preferably with a non-toxic molecule that
could be given orally to cancer patients,”
enthuses Hellmich. However, this is not a
straightforward endeavour as CBS activity does
not only occur in cancerous cells but in other
tissues and organs where it has important
signalling functions. Therefore, the researchers
are currently directing their studies toward
identifying a safe limit of CBS inhibition that
disrupts its cancer-promoting activity without
affecting the normal H2S-associated signalling
pathways of other normal body tissues.
POTENTIAL AVENUES
FOR FURTHER RESEARCH
Although the researchers’ experiments to
date have enlightened them on the three
ways in which CBS activity promotes cancer
cell growth, the mechanisms by which these
occur remain elusive. The collaborators are
actively seeking funding to embark upon this
line of investigation, as well as other pertinent
research. For example, the phenomena whereby
dogs are able to identify patients with cancer
based on their smell is relatively well-known.
The finding that the strong-smelling H2S
gas is elevated in the tumour cells of cancer
patients could explain this long-questioned
peculiarity. Over the last few years, research
published by other research groups has been
developing highly sensitive cancer diagnostic
strategies that involve detecting increased H2S
levels in the urine or breath of cancer patients.
This is another area of research that Szabo
and Hellmich would like to explore – tools to
detect elevated levels of H2S that would enable
appropriate patients to be matched up with
CBS inhibitory therapies.
Armed with the knowledge that CBSsynthesised H2S in cancer cells is essential for
energy production to enable their division,
growth and invasion of host tissue, Szabo and
Hellmich have founded CBS Therapeutics, a
startup company involved in research and
development of CBS inhibitors to target colon
cancer. “We are actively working on raising
additional funds, both for future basic research
and for the commercialisation and therapeutic
translation of our concept for patient benefit,”
Szabo concludes. While the collaborators
recognise that these are very early days in the
field of H2S and cancer biology, they envision
various accelerated paths for the successful
clinical translation of their discoveries.
CONTACT
Dr Csaba Szabo
Professor of Anaesthesiology
Department of Anesthesiology
University of Texas Medical Branch
Room 4.2.2H, Building 21
610 Harborside Drive
Galveston Texas 77555-1102, USA
T +1 206 291 2959
E [email protected]
DR CSABA SZABO is a tenured Professor
of Anesthesiology at the University of Texas
Medical Branch, Galveston, Texas, USA. He
received his MD and PhD (Physiology) at
the Semmelweis University Medical School,
Budapest, Hungary, and holds a second PhD
(Pharmacology) from the University of London,
UK. Szabo’s research interests include the
pathomechanisms of various forms of critical
illness, the molecular mechanisms of cell death,
mitochondrial dysfunction, vascular injury, free
radicals biology, poly(ADP-ribose) polymerase,
nitric oxide and hydrogen sulphide. He is a
recognised expert in the field of hydrogen
sulphide; he has identified novel vascular and
metabolic roles of this mediator in angiogenesis,
circulatory shock, heart disease and diabetes,
and, most recently, in collaboration with Dr
Hellmich, in gastrointestinal cancer.
DR MARK HELLMICH is a tenured Professor in
the Departments of Surgery and Neuroscience
and Cell Biology. He holds a PhD (Physiology)
from Boston University and completed
postdoctoral training at the National Cancer
Institute. Hellmich is a recognised expert in
the molecular pathophysiology of human
gastrointestinal malignancies. His interests
include: 1) the role of peptide hormones
and their receptors in regulating the
interactions between cancer cells and tumour
microenvironment; 2) tumour stromal cellspecific markers that predict the recurrence
of disease; and 3) molecular mechanisms
involved in bone marrow cell differentiation and
activation. His prior achievements include the
first discovery and characterisation of a neuronal
cdc2-like kinase later called CDK5 and isolation
of the first enzyme to produce the calciummobilising mediator cyclic ADP-ribose.
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