Journal club presentation Ying-Hua Wang et. al. 2014 Cell Cell-State-Specific Metabolic Dependency in Hematopoiesis and Leukemogenesis

1. Journal club
9.2.15

2. Pyruvate Kinase (PK) and Lactate Dehydrogenase
(LDH) Represent a Potential Anticancer Targets
• PK catalyzes the conversion of
phosphoenolpyruvate (PEP) and ADP
to pyruvate and ATP.
• PKM1 is expressed in differentiated
adult tissues that have a high demand
for ATP production and metabolize
glucose preferentially via oxidative
phosphorylation.
• PKM2 is expressed in early embryonic
tissues, cancers, and adult cells that
have high anabolic activity.

3. The Authors Question
• How does modulating glucose metabolism
specifically affect hematopoietic and leukemic
cell populations?
Glucose
metabolism
HSCs
Progenitor
cells
Leukemic
cells
PKM2 KO LDHA KO

4. PKM2 IS THE PREDOMINANT ISOFORM OF PK EXPRESSED IN NORMAL
HEMATOPOIETIC AND LEUKEMIC CELLS
CONDITIONAL DELETION OF PKM2 IN THE HEMATOPOIETIC LINEAGE
Taken together, these results suggest
that incomplete repression of exon 9
during splicing allows some PKM1
expression in hematopoietic cells
following excision of exon 10 but that
overall PK expression is less than that
in WT cells.

5. DELETION OF PKM2 DOES NOT AFFECT HEMATOPOIESIS UNDER
HOMEOSTATIC CONDITIONS BUT COMPROMISES LONG-TERM
HEMATOPOIESIS IN TRANSPLANTATION
The competitive disadvantage of
PKM2−/− cells in the BM was
observed in progenitor
populations.
Pkm2−/− HSPCs displayed a
marked defect in repopulating
both myeloid and lymphoid
progeny as early as 4–8 weeks
following transplantation.
PKM2 deletion impairs the BM
repopulating capacity cell-
autonomously, likely by affecting
progenitor cell proliferation.

6. METABOLIC CHANGES INDUCED BY PKM2 ABLATION AND PKM1 EXPRESSION
Oxygen concentrations in
the BM environment are
closer to 1%.
TCA Cycle
Taken together, these data are consistent
with the hypothesis that PKM2 deletion with
PKM1 expression impairs AG, enhances
oxidative metabolism, and limits the
macromolecule biosynthesis required for cell
proliferation.

7. Deletion of PKM2 in BM cells
 Had no effect on the maintenance of normal haematopoiesis.
 At competitive BM repopulation assays progenitor cells lacking PKM2 (but not
long-term HSCs) had a reduced ability to reconstitute mature blood cells.
 Haematopoietic progenitor cells lacking PKM2 had increased oxidative
metabolism, increased mitochondrial membrane potential and decreased lactate
production.
 The authors also showed that cells that lacked PKM2 then expressed PKM1; as
PKM1 promotes oxidative phosphorylation, this is consistent with the metabolic
changes observed in these cells.

8. DELETION OF LDHA COMPROMISES LONG-TERM BM REPOPULATION CAPACITY
VIA A DIFFERENT MECHANISM THAN THAT OBSERVED IN PKM2−/− MICE
•The percentage of
cycling cells (S/G2/M)
was lower in both
progenitor and stem
populations
from Ldha−/− mice
•Secondary
transplantation
•Primary transplantation
•LDHA is predominant
isoform in HSPC
•Loss of LDHA abrogated
both aerobic and
anaerobic glycolysis

9. Both basal and maximal
respiration rates were elevated
markedly in Ldha−/− Lin− cells
compared with WT control
cells.
The functional rescue
of Ldha−/− cells by NAC appears to
be at the level of stem cells because
the long-term HSCs in the BM were
partially recovered by NAC
DELETION OF LDHA COMPROMISES LONG-TERM BM REPOPULATION CAPACITY
VIA A DIFFERENT MECHANISM THAN THAT OBSERVED IN PKM2−/− MICE Cont.

10. • Progenitor cells lacking LDHA had defects in BM repopulation.
• But in contrast to the results with PKM2, LDHA loss reduced the
repopulating ability of HSCs as well.
• Oxidative state and mitochondrial membrane potential were also
increased in LDHA-null cells.
• LDHA-null cells had increased levels of reactive oxygen species
(ROS), which was not observed in cells lacking PKM2.
Deletion of LDHA in BM cells

12. Chronic myeloid leukemia (CML) Acute myeloid leukemia (AML)
BCR-ABL MLL-AF9
stem cell-based disease myeloid progenitor (GMP) disease
PKM2 KO
LDHA KO
Control
5-FU
injection
5-6 days BM harvesting and
infection with retrovirus
Injection with normal
supporting BM cells
into lethally irradiated
mice

13. IMPACT OF ALTERED GLYCOLYSIS ON LEUKEMOGENESIS IN VIVO
For both CML and AML, mice transplanted
with Pkm2−/− or Ldha−/− cells displayed
significantly prolonged disease latency.
Depletion of PKM2 or LDHA enforces a
metabolic shift from glycolysis to
mitochondrial respiration and
compromises leukemia induction in
mice.

14. Initiation vs. Maintenance

15. Summary
• Knockout of both proteins suppressed leukaemia progression but
did not affect normal haematopoietic cells.
• Although neither of these enzymes is absolutely required for
leukaemogenesis, leukaemia cells are more sensitive to glycolysis
inhibition than normal cells.
• A therapeutic window might exist in which moderate suppression
of glycolysis could prevent leukaemia progression but spare normal
haematopoietic stem and progenitor cells.