title: Small-molecule inhibitors of Ataxia Telangiectasia and Rad3 related kinase (ATR) sensitize lymphoma cells to UVA radiation.
abstract: BACKGROUND
  Psoralen plus ultraviolet A (PUVA) photochemotherapy is a combination treatment used for inflammatory and neoplastic skin diseases such as mycosis fungoides (MF), the most common type of cutaneous T-cell lymphoma (CTCL). However, 30% of MF patients do not respond sufficiently to PUVA and require more aggressive therapies.
  
  
  OBJECTIVE
  The aim of this project was to investigate whether inhibition of Ataxia Telangiectasia and Rad3 related kinase (ATR) may enhance efficacy of phototherapy.
  
  
  METHODS
  CTCL cell lines (MyLa2000, SeAx and Mac2a) served as in vitro cell models. ATR and Chk1 were inhibited by small molecule antagonists VE-821, VE-822 or Chir-124, or by small interfering RNAs (siRNAs). Cell cycle and viability were assessed by flow cytometry.
  
  
  RESULTS
  Small molecule inhibitors of ATR and Chk1 potently sensitized all cell lines to PUVA and, importantly, also to UVA, which by itself did not cause apoptotic response. VE-821/2 blocked ATR pathway activation and released the cells from the G2/M block caused by UVA and PUVA, but did not affect apoptosis caused by other chemotherapeutics (etoposide, gemcitabine, doxorubicine) or by hydrogen peroxide. Knockdown of ATR and Chk1 with siRNA also blocked the ATR pathway and released the cells from G2/M block but did not sensitize the cells to UVA as observed with the small molecule inhibitors. The latter suggested that the synergism between VE-821/2 or Chir-124 and UVA was not solely caused by specific blocking of ATR kinase but also ATR-independent photosensitization. This hypothesis was further verified by administrating VE-821/2 or Chir-124 before and after UVA irradiation, as well as comparing their activity with other ATR and Chk1 inhibitors (AZD6738 and MK8776). We found that only VE-821/2 and Chir-124 kinase inhibitors had synergistic effect with UVA, and only if applied before treatment with UVA.
  
  
  CONCLUSIONS
  Small molecule ATR and Chk1 inhibitors potently sensitize lymphoma cells to UVA radiation and induce a prominent apoptotic response. Interestingly, this effect is due to the dual (kinase inhibiting and photosensitizing) mode of action of these compounds.
authors.affiliation: Department of Dermatology, Bispebjerg Hospital, Copenhagen, Denmark. Electronic address: edyta.biskup@gmail.com.
authors.forename: Edyta
authors.initials: E
authors.lastname: Biskup
authors.affiliation: Department of Dermatology, Bispebjerg Hospital, Copenhagen, Denmark.
authors.forename: David Gram
authors.initials: DG
authors.lastname: Naym
authors.affiliation: Department of Dermatology, Bispebjerg Hospital, Copenhagen, Denmark; Faculty of Health Sciences, University of Copenhagen, Denmark; Division of Dermatology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Canada.
authors.forename: Robert
authors.initials: R
authors.lastname: Gniadecki
mesh_terms: D064007:Ataxia Telangiectasia Mutated Proteins / Q000037:antagonists & inhibitors; D002453:Cell Cycle; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002470:Cell Survival; D000071877:Checkpoint Kinase 1 / Q000037:antagonists & inhibitors*; D005434:Flow Cytometry; D006657:Histones / Q000378:metabolism; D006801:Humans; D007555:Isoxazoles / Q000494:pharmacology; D016410:Lymphoma, T-Cell, Cutaneous / Q000378:metabolism / Q000473:pathology* / Q000628:therapy; D010766:Phosphorylation; D010778:Photochemotherapy; D017319:Photosensitizing Agents / Q000494:pharmacology*; D011719:Pyrazines / Q000494:pharmacology; D011804:Quinolines / Q000494:pharmacology; D011812:Quinuclidines / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000378:metabolism; D013450:Sulfones / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D014466:Ultraviolet Rays
chemical_list: C560580:3-amino-6-(4-(methylsulfonyl)phenyl)-N-phenylpyrazine-2-carboxamide; C518919:CHIR-124; C468783:H2AX protein, human; D006657:Histones; D007555:Isoxazoles; D017319:Photosensitizing Agents; D011719:Pyrazines; D011804:Quinolines; D011812:Quinuclidines; D034741:RNA, Small Interfering; D013450:Sulfones; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; C000598331:berzosertib
keywords: ATR inhibitors; Cutaneous T-cell lymphoma; DNA damage; VE-821; VE-822