The Novel Anti-Cancer Agent, SpiD3, Is Cytotoxic in CLL Cells Resistant to Ibrutinib or Venetoclax
, Audrey L. Smith 1, Sydney A. Skupa 1, Kabhilan Mohan 1, Sandeep Rana 1, Sarbjit Singh 1, Jayapal Reddy Mallareddy 1, Grinu Mathew 1,2, Amarnath Natarajan 1,2 and Dalia El-Gamal 1,2,*
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsThank you for this interesting paper evaluating SpiD3 in ibrutinib or venetoclax in resistance in chronic lymphocytic leukemia models.
There are some background comments that could allow for context and importance of this work.
1. How were the ibr and ven resistance created in the osu cell lines ( did you develop or were they provided?)
2. More backgrpund on ferroptosis should be included and relevance to other b cell malignancies and context for treatment should be added.
3. The combination of BCL 2 and BTKI is sweeping the practice by storm. None of the experiments evaluate :
a. efficacy or mechanism in double refractory models which will be key. As we have to understand the position of this agent.
b. What does the ven ibr combo vs SpiD3 look like also what is the comparison between bcl2 bv btk vs the combo. This is essential in further understanding the roleof ferroptosis and potential mechanisms.
4. The western blots are beautiful but I don’t see information on replicates , bleed demonstrate validation across 2-3 experiments and provide a graphical representation. Also the molecular profile of the cell line is not described also what is the resistance mechanism and is it generalizable. Many patients would have a p53 mutation historically but withless chemotherapy being used is this a relevant model? Please replicate in other b cell cll models.
5. Thapsigargin. Some would state that thapsi alters the notch signalling pathway why what is its role with redox pathway and the suitability for this experiment.
6. There is significant PARP cleavage, what is happening to caspase cleavage? What is happening to mitochondrial function and respiration ? is this molecule affecting complex 1 activity? Is Gamma gammaH2AX altered?
Author Response
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Author Response File: 
Author Response.pdf
Reviewer 2 Report
Comments and Suggestions for AuthorsAlexandria P Eiken, et al, report a very interesting manuscript, describing a new anti-cancer agent, SpiD3, in ibrutinib or venetoclax resistant CLL cells. They also performed many and adequate methods to find out the mechanisms of overcome resistance pathways and prove them. There are 2 things need to be clarified.
1. In "discussion" section, Supplementary Figure S2D, and S4D, you mentioned that the lower molecular weight band of SLC3A2 observed in immunoblot analysis of SpiD3-treated cells, idicates a monomer form of SLC3A2 to interfere with cystine impport. Would you please provide the ladder picture to let us know what the molecular weight the band is?
2. I found one article " A Novel Spirocyclic Dimer (SpiD3) Resensitizes CLL cells to venetoclax", published from your group, on Blood (2022) 140 (Supplement 1): 8872-3. The Figure is very similar to your Figure 4B. Please cite this reference.
Overall, this is a good paper.
Author Response
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Author Response File: Author Response.pdf
