Reposted by Hooman HefziNathan Lewis @natelewis3.bsky.social · 03/02/2025Took me a while to get the new uga address, but my lab website is finally live. lewislab.uga.edu 162
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025I am also really glad that people far more creative than I dressed up a hamster as Godzilla so that when I decided to name the cell line CHO ZeLa (for Zero Lacate) I could use lots of fun pictures for presentations. 010
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Finally, this work could not have been done without the tireless work of 30+ other authors from 7+ institutions over ~8 years! Special shout outs to @natelewis3.bsky.social and Bjørn Voldborg for humoring me when I asked them to knock out these 5 genes at once just to give it a shot. 120
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025I've recently started as an Associate Professor at the Technical University of Denmark and my new research group is definitely interested in poking at this in more detail :) It's a big space so reach out if you think a collaboration could be fun! 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Sosososo many interesting future lines of inquiry. I can't say we've figured out the "why" of the Warburg effect, but I think it is safe to say that it is definitely dispensable for proliferation. 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Strikingly, compared to a human Warburg-null LS174T cancer line (LDHA/B double KO with reduced growth), we saw almost no overlap in consistently, differentially expressed genes (vs WT); in fact, more genes showed consistent opposite directionality in CHO Warburg-null cells. 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Metabolic genes linked to redox metabolism, however, did seem generally perturbed. 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025When we looked at RNA-Seq to identify an underlying basis for the Warburg-null phenotype, we were surprised to see very little overlap in differentially expressed genes (vs WT) when looking at Warburg-null clones derived from 2 different CHO cell lines. 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Unsurprisingly, Warburg-null cells: • Used slightly more oxygen • Showed increased glucose-->TCA cycle flux • Were more sensitive to mitochondrial inhibitors • Showed a significantly lower NAD/NADH ratio 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025We confirmed that the Warburg-null CHO cells could still be used to make biotherapeutic proteins in high quantity and quality and turned to investigate how Warburg-null cells responded to elimination of this ubiquitous phenotype. 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025We later showed that this same approach was effective in another CHO cell line as well as HEK293, making us think that this may be a generalizable genetic engineering strategy to eliminate the Warburg effect! 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025We later made a range of knockouts and showed that knocking out even 1 (of 4) pyruvate dehydrogenase kinases enabled the knockout of Ldha and elimination of the Warburg effect. These cells grew as well as wildtype, did not produce lactate, and consumed much less glucose. 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025We first generated knockouts of all pyruvate dehydrogenase kinases and Ldha. We confirmed that lactate dehydrogenase is not detected, that cells do not produce lactate, and show negligible lactate dehydrogenase activity (wildtype in blue, knockout lines in red). 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025~8(!) years ago, we tried to eliminate the Warburg effect in CHO cells and found that simultaneous knock out of pyruvate dehydrogenase kinases and lactate dehydrogenases eliminates lactate production w/out impacting growth! 110
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Thus, significant effort has been dedicated to eliminate lactate production in these lines, but none have proven effective. This is consistent with observations that targeting lactate production in cancer leads to cell death, though this has not led to approved therapeutics. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Mammalian lines such as Chinese hamster ovary (CHO) cells and HEK293 are broadly used for the production of life-saving biotherapeutics, and also exhibit the Warburg effect. Lactate accumulation in cell culture is detrimental to growth and, thus, biotherapeutic production. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Over the years, there has been an incredible amount of research trying to answer the "why" of this phenomenon. Some theorized that it was necessary for rapid ATP generation or to provide the necessary precursors for biosynthetic pathways. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025101 years ago, Otto Warburg observed that cancer cells produced high quantities of lactate from glucose, even in the presence of oxygen. Later research suggested that this "Warburg effect" was a hallmark of all proliferative mammalian cells, e.g., stem cells. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025After ~8 years, happy to share work done w/ @natelewis3.bsky.social showing you can eliminate the Warburg effect in mammalian cells w/out impacting growth rate is out now in @natmetabolism.bsky.social! www.nature.com/articles/s42... #science #biotech #cancermetabolism Thread below:nature.comMultiplex genome editing eliminates lactate production without impacting growth rate in mammalian cells - Nature MetabolismHefzi et al. engineer cells to nearly eliminate lactate production and increase mitochondrial use of pyruvate by deleting both lactate dehydrogenases and pyruvate dehydrogenase kinases with the goal o... 1178
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025I am also really glad that people far more creative than I dressed up a hamster as Godzilla so that when I decided to name the cell line CHO ZeLa (for Zero Lacate) I could use lots of fun pictures for presentations. 000
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Finally, this work could not have been done without the tireless work of 30+ other authors from 7+ institutions over ~8 years! Special shout outs to @nate3791.bsky.social and Bjørn Voldborg for humoring me when I asked them to knock out these 5 genes at once just to give it a shot. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025I've recently started as an Associate Professor at the Technical University in Denmark and my new research group is definitely interested in poking at this in more detail :) It's a big space so reach out if you think a collaboration could be fun! 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Sosososo many interesting future lines of inquiry. I can't say we've figured out the "why" of the Warburg effect, but I think it is safe to say that it is definitely dispensable for proliferation. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Strikingly, compared to a human Warburg-null LS174T cancer line (LDHA/B double KO with reduced growth), we saw almost no overlap in consistently, differentially expressed genes (vs WT); in fact, more genes showed consistent opposite directionality in CHO Warburg-null cells. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Metabolic genes linked to redox metabolism, however, did seem generally perturbed. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025When we looked at RNA-Seq to identify an underlying basis for the Warburg-null phenotype, we were surprised to see very little overlap in differentially expressed genes (vs WT) when looking at Warburg-null clones derived from 2 different CHO cell lines. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Unsurprisingly, Warburg-null cells: • Used slightly more oxygen • Showed increased glucose-->TCA cycle flux • Were more sensitive to mitochondrial inhibitors • Showed a significantly lower NAD/NADH ratio 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025We confirmed that the Warburg-null CHO cells could still be used to make biotherapeutic proteins in high quantity and quality and turned to investigate how Warburg-null cells responded to elimination of this ubiquitous phenotype. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025We later showed that this same approach was effective in another CHO cell line as well as HEK293, making us think that this may be a generalizable genetic engineering strategy to eliminate the Warburg effect! 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Finally, we grew cells with Ldha and 1 (purple), 2 (green), 3 (orange), and 4 (red) pyruvate dehydrogenase kinases knocked out for ~3 months and showed that the Warburg-null phenotype was stable over time. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025We later made a range of knockouts and showed that knocking out even 1 (of 4) pyruvate dehydrogenase kinases enabled the knockout of Ldha and elimination of the Warburg effect. These cells grew as well as wildtype, did not produce lactate, and consumed much less glucose. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025We first generated knockouts of all pyruvate dehydrogenase kinases and Ldha. We confirmed that lactate dehydrogenase is not detected, that cells do not produce lactate, and show negligible lactate dehydrogenase activity (wildtype in blue, knockout lines in red). 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025~8(!) years ago, we tried to eliminate the Warburg effect in CHO cells and found that simultaneous knock out of pyruvate dehydrogenase kinases and lactate dehydrogenases eliminates lactate production w/out impacting growth! 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Mammalian lines such as Chinese hamster ovary (CHO) cells and HEK293 are broadly used for the production of life-saving biotherapeutics, and also exhibit the Warburg effect. Lactate accumulation in cell culture is detrimental to growth and, thus, biotherapeutic production. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025Over the years, there has been an incredible amount of research trying to answer the "why" of this phenomenon. Some theorized that it was necessary for rapid ATP generation or to provide the necessary precursors for biosynthetic pathways. 100
Hooman Hefzi @hefzi-lab.bsky.social · 20/01/2025101 years ago, Otto Warburg observed that cancer cells produced high quantities of lactate from glucose, even in the presence of oxygen. Later research suggested that this "Warburg effect" was a hallmark of all proliferative mammalian cells, e.g., stem cells. 100
