Selective elimination of pluripotent stem cells by PIKfyve specific inhibitors

Chakraborty, Arup; Vassilev, Alex; Jaiswal, Sushila; O'Connell, Constandina E.; Ahrens, John F.; Mallon, Barbara S.; Pera, Martín F.; DePamphilis, Melvin L.

doi:10.1016/j.stemcr.2021.12.013

Cited by 6 publications

(25 citation statements)

References 58 publications

(103 reference statements)

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“…As previously reported, PIKFYVE inhibitors induce noncanonical apoptosis selectively in PIKFYVEsensitive cells [4,7,9,13]. Loss of DNA in A375 cells was 709 more sensitive to WX8 than HFF1 cells (Fig.…”

Section: Il24 Was Essential For Wx8 Induction Of Cell Deathsupporting

confidence: 71%

“…Examples include lymphoma [3,4], Abbreviations Adr, Adriamycin; Apl, apilimod; BafA1, bafilomycin-A1; CQ, chloroquine; DMSO, dimethyl sulfoxide; ELISA, enzyme-linked immunosorbent assay; ER, endoplasmic reticulum; Etop, etoposide; FACS, fluoresce activated cell sorting; IL24, interleukin-24; ns, not significant; nt-siRNA, nontarget siRNA; PCR, polymerase chain reaction; pfu, plaque forming units; PI(3,5)P 2 , phosphatidyl inositol 3,5-bisphosphate; PI(4)P, phosphatidyl inositol 4-phosphate; PI(4,5)P 2 , phosphatidyl inositol 4,5-bisphosphate; PI(5)P, phosphatidyl inositol 5-phosphate; PIKFYVE, 1-phosphatidylinositol 3-phosphate 5-kinase activity; PIP4K2C, 1-phosphatidylinositol 5-phosphate 4-kinase activity; PIP5K1C, 1-phosphatidylinositol 4-phosphate 5-kinase activity; RNA-seq, RNA sequence profiling; RT-PCR, real-time polymerase chain reaction; SD, standard deviation; SEM, standard error of the mean; STR, short tandem repeat; THAP, thapsigargin; TUN, tunicamycin; V, vehicle (DMSO); Vac, Vacuolin-1; WT, wild-type; YM, YM201636. liver [5], multiple myeloma [6], colorectal [7], prostate [8], embryonal carcinoma [9], gastric [10], and melanoma (this report). Thus, PIKFYVE inhibitors can selectively terminate autophagy-dependent cancer cells and pluripotent cancer stem cells under conditions where nonmalignant cells remain viable [4,7,9,[11][12][13][14].…”

Section: Introductionmentioning

confidence: 74%

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PIKFYVE inhibitors trigger interleukin‐24‐dependent cell death of autophagy‐dependent melanoma

Roy,

Chakraborty,

DePamphilis

2024

Molecular Oncology

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Inhibitors specifically targeting the 1‐phosphatidylinositol 3‐phosphate 5‐kinase (PIKFYVE) disrupt lysosome homeostasis, thereby selectively terminating autophagy‐dependent human cancer cells in vivo as well as in vitro without harming the viability of nonmalignant cells. To elucidate the mechanism by which PIKFYVE inhibition induces cell death, autophagy‐dependent melanoma cells were compared with normal foreskin fibroblasts. RNA sequence profiling suggested that PIKFYVE inhibitors upregulated an endoplasmic reticulum (ER) stress response involving interleukin‐24 (IL24; also known as MDA7) selectively in melanoma cells. Subsequent biochemical and genetic analyses confirmed these results and extended them to tumor xenografts in which tumor formation and expansion were inhibited. IL24 expression was upregulated by the DDIT3/CHOP/CEBPz transcription factor, a component of the PERK‐dependent ER‐stress response. Ectopic expression of IL24‐induced cell death in melanoma cells, but not in foreskin fibroblasts, whereas ablation of the IL24 gene in melanoma cells prevented death. IL24 upregulation was triggered specifically by PIKFYVE inhibition. Thus, unlike thapsigargin and tunicamycin, which induce ER‐stress indiscriminately, PIKFYVE inhibitors selectively terminated PIKFYVE‐sensitive melanoma by inducing IL24‐dependent ER‐stress. Moreover, induction of cell death by a PIKFYVE inhibitor together with ectopic expression of IL24 protein was cumulative, thereby confirming the therapeutic potential of PIKFYVE inhibitors in the treatment of melanoma.

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Section: Il24 Was Essential For Wx8 Induction Of Cell Deathsupporting

confidence: 71%

Section: Introductionmentioning

confidence: 74%

PIKFYVE inhibitors trigger interleukin‐24‐dependent cell death of autophagy‐dependent melanoma

Roy,

Chakraborty,

DePamphilis

2024

Molecular Oncology

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“…The crucial components to be evaluated for the efficient gene editing of PSC are the choice of the delivery vectors and of the suicide genes with their relative selection marker, as the selection of edited cells is fundamental to obtain a pure edited population. Selection methods include the addition of antibioticresistant cassettes or genome-edited cell sorting based on the induced expression of fluorescent reporters or surface antigens (90)(91)(92). These selection strategies can also be combined to obtain a purer population or to select a cell population edited with more than one construct.…”

Section: A Safety Switch For a Safer Cell Therapymentioning

confidence: 99%

“…Molecules capable of targeting various other elements of the cell, such as lysosomes, proteins, and pumps present on the cell membrane, have also been described. Recently, Chakraborty et al explored the use of WX8 and apilimod as inhibitors of PIKfyve phosphatidylinositol kinase, which is essential for lysosome homeostasis, to selectively kill PSC under conditions where differentiated cells remain viable ( 91 ). PIKfyve inhibitors prevent lysosome fission, induce autophagosome accumulation, and reduce cell proliferation in both pluripotent and differentiated cells, but induce death specifically in pluripotent cells by non-canonical apoptosis ( 91 ).…”

Section: Depletion Of Contaminant Pluripotent Stem Cells In the Final...mentioning

confidence: 99%

“…Recently, Chakraborty et al explored the use of WX8 and apilimod as inhibitors of PIKfyve phosphatidylinositol kinase, which is essential for lysosome homeostasis, to selectively kill PSC under conditions where differentiated cells remain viable ( 91 ). PIKfyve inhibitors prevent lysosome fission, induce autophagosome accumulation, and reduce cell proliferation in both pluripotent and differentiated cells, but induce death specifically in pluripotent cells by non-canonical apoptosis ( 91 ). Recently, it has been shown that bee venom (BV) can specifically induce cell death in iPSC but not in iPSC-derived differentiated cells; however, the cause of this selectivity has yet to be clarified.…”

Section: Depletion Of Contaminant Pluripotent Stem Cells In the Final...mentioning

confidence: 99%

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Strategies to Improve the Safety of iPSC-Derived β Cells for β Cell Replacement in Diabetes

Pellegrini¹,

Zamarian²,

Sordi³

2022

Transpl Int

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Allogeneic islet transplantation allows for the re-establishment of glycemic control with the possibility of insulin independence, but is severely limited by the scarcity of organ donors. However, a new source of insulin-producing cells could enable the widespread use of cell therapy for diabetes treatment. Recent breakthroughs in stem cell biology, particularly pluripotent stem cell (PSC) techniques, have highlighted the therapeutic potential of stem cells in regenerative medicine. An understanding of the stages that regulate β cell development has led to the establishment of protocols for PSC differentiation into β cells, and PSC-derived β cells are appearing in the first pioneering clinical trials. However, the safety of the final product prior to implantation remains crucial. Although PSC differentiate into functional β cells in vitro, not all cells complete differentiation, and a fraction remain undifferentiated and at risk of teratoma formation upon transplantation. A single case of stem cell-derived tumors may set the field back years. Thus, this review discusses four approaches to increase the safety of PSC-derived β cells: reprogramming of somatic cells into induced PSC, selection of pure differentiated pancreatic cells, depletion of contaminant PSC in the final cell product, and control or destruction of tumorigenic cells with engineered suicide genes.

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PIP5K1C phosphoinositide kinase deficiency distinguishes PIKFYVE-dependent cancer cells from non-malignant cells

Chakraborty

Nomanbhoy²,

DePamphilis

2023

Autophagy

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Although PIKFYVE phosphoinositide kinase inhibitors can selectively eliminate PIKFYVE-dependent human cancer cells in vitro and in vivo , the basis for this selectivity has remained elusive. Here we show that the sensitivity of cells to the PIKFYVE inhibitor WX8 is not linked to PIKFYVE expression, macroautophagic/autophagic flux, the BRAF V600E mutation, or ambiguous inhibitor specificity. PIKFYVE dependence results from a deficiency in the PIP5K1C phosphoinositide kinase, an enzyme required for conversion of phosphatidylinositol-4-phosphate (PtdIns4P) into phosphatidylinositol-4,5-bisphosphate (PtdIns[4,5]P 2 /PIP2), a phosphoinositide associated with lysosome homeostasis, endosome trafficking, and autophagy. PtdIns(4,5)P 2 is produced via two independent pathways. One requires PIP5K1C; the other requires PIKFYVE and PIP4K2C to convert PtdIns3P into PtdIns(4,5)P 2 . In PIKFYVE-dependent cells, low concentrations of WX8 specifically inhibit PIKFYVE in situ , thereby increasing the level of its substrate PtdIns3P while suppressing PtdIns(4,5)P 2 synthesis and inhibiting lysosome function and cell proliferation. At higher concentrations, WX8 inhibits both PIKFYVE and PIP4K2C in situ , which amplifies these effects to further disrupt autophagy and induce cell death. WX8 did not alter PtdIns4P levels. Consequently, inhibition of PIP5K1C in WX8-resistant cells transformed them into sensitive cells, and overexpression of PIP5K1C in WX8-sensitive cells increased their resistance to WX8. This discovery suggests that PIKFYVE-dependent cancers could be identified clinically by low levels of PIP5K1C and treated with PIKFYVE inhibitors. Abbreviations: DMSO: dimethylsulfoxide; ELISA: enzyme-linked immunosorbent assay; LC3-I: microtubule associated protein light chain 3-I; LC3-II: microtubule associated protein light chain 3-II; MS: mass spectrometry; PtdIns: phosphatidylinositol; PtdIns3P: PtdIns-3-phosphate; PtdIns4P: PtdIns-4-phosphate; PtdIns5P: PtdIns-5-phosphate; PtdIns(3,5)P 2 : PtdIns-3,5-bisphosphate; PtdIns(4,5)P 2 /PIP2: PtdIns-4,5-bisphosphate; PtdIns(3,4,5)P 3 /PIP3: PtdIns-3,4,5-trisphosphate; PIKFYVE: phosphoinositide kinase, FYVE-type zinc finger containing; PIK3C3: phosphatidylinositol 3-kinase catalytic subunit type 3; PI4KA: phosphatidylinositol 4-kinase alpha; PI4KB: phosphatidylinositol 4-kinase beta; PI4K2A: phosphatidylinositol 4-kinase type 2 alpha; PI4K2B: phosphatidylinositol 4-kinase type 2 beta; PIP4K2A: phosphatidylinositol-5-phosphate 4-kinase type 2 alpha; PIP4K2B: phosphatidylinositol-5-phosphate 4-kinase type 2 beta; PIP4K2C: phosphatidylinositol-5-phosphate 4-kinase type 2 gamma; PIP5K1A: phosphatidylinositol-4-phosphate 5-kinase type 1 alpha; PIP5K1B: phosphatidylinositol...

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Selective elimination of pluripotent stem cells by PIKfyve specific inhibitors

Cited by 6 publications

References 58 publications

PIKFYVE inhibitors trigger interleukin‐24‐dependent cell death of autophagy‐dependent melanoma

PIKFYVE inhibitors trigger interleukin‐24‐dependent cell death of autophagy‐dependent melanoma

Strategies to Improve the Safety of iPSC-Derived β Cells for β Cell Replacement in Diabetes

PIP5K1C phosphoinositide kinase deficiency distinguishes PIKFYVE-dependent cancer cells from non-malignant cells

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