Synthetic Lethality Approaches in Acute Lymphoblastic Leukemia

Round 1

Reviewer 1 Report

Lagunas-Rangel and Chavez-Valencia provide a comprehensive review on synthetic lethality (SL) and treatment of Acute Lymphoblastic Leukemia (ALL).

The review covers the few previously published cases where SL treatment has shown promise and attempts to discuss potential novel targets that could be exploited for treatment.  There is a good introduction which helps to explain to the concept to non-experts in the leukemia field, but there are sections within the article that read like a list of findings and the organization of these could be strengthened – or shortened.  

SL has not so far been particularly forthcoming in the treatment of leukemia – it would be more interesting to know whether any of the many combinations presented in the article have been, or are to be, tested on patients?

ALL arises in B-lymphocytes in the early stages of development in the bone marrow. The disease is usually therefore called precursor B-cell ALL or pre-B cell ALL.  B-cell ALL arises in more mature developing lymphocytes.  

In Table 1, the authors should really distinguish between BCP-ALL and B-ALL in the different leukemia subtypes.

Although Table 1 is quite informative for the subtypes, the review itself overly concentrates on cases of BCR-ABL1+ ALL or T-cell ALL.   The authors could discuss the other leukemia fusion gene subtypes mentioned – even only as potential candidates for SL treatment? 

For example, epigenetic SL has been proposed for MLL-r acute leukemias (mainly in AML), but PARP inhibitors, HDAC inhibitors etc may all be able to play a role in BCP-ALL treatment through SL and should be discussed in some more detail.

For Table 2, perhaps the title should also state "in ALL cell lines"?

In Table 2 it is quite difficult to interpret which of the genes shown to be involved in the different leukemia subtypes correspond to the observations detected after SL treatment.  Better spacing of the lines between the subtypes in the table may help?   Why is BCR-ABL1 fusion only shown for one Ph+ subtype?  Should the relevant TKI’s or other used inhibitors also be shown in the Table?

Minor:

Line 55: What is the percentage of overall relapse (what does high incidence mean here?).  Isn't leukemia relapse sub-type dependent?

Author Response

Dear Reviewer

We thank you for taking the time to review our work and for your valuable comments, which help us to improve its quality. Responses to their comments are given below, and all modifications to the manuscript are indicated in red text.

Reviewer 1

Lagunas-Rangel and Chavez-Valencia provide a comprehensive review on synthetic lethality (SL) and treatment of Acute Lymphoblastic Leukemia (ALL).

The review covers the few previously published cases where SL treatment has shown promise and attempts to discuss potential novel targets that could be exploited for treatment.  There is a good introduction which helps to explain to the concept to non-experts in the leukemia field, but there are sections within the article that read like a list of findings and the organization of these could be strengthened – or shortened.  

SL has not so far been particularly forthcoming in the treatment of leukemia – it would be more interesting to know whether any of the many combinations presented in the article have been, or are to be, tested on patients?

R1.

Based on your feedback, we incorporated a sentence highlighting its application in patients. In the manuscript, this addition reads as follows:

 For these reasons, unfortunately only one synthetic lethal interaction –that between PARP and BRCA1 and/or BRCA2– has successfully moved from discovery to clinical approval. Specifically in the ALL context, it is noteworthy that most synthetic lethality findings, so far, have been identified only in cell lines, with a limited number in murine models. For this reason, ALL treatment landscape lacks ongoing clinical trials dedicated to exploring synthetic lethality strategies. In this sense, to change this trend, it is imperative to broaden the scope of research and address the associated challenges through interdisciplinary collaboration.

ALL arises in B-lymphocytes in the early stages of development in the bone marrow. The disease is usually therefore called precursor B-cell ALL or pre-B cell ALL.  B-cell ALL arises in more mature developing lymphocytes.  

In Table 1, the authors should really distinguish between BCP-ALL and B-ALL in the different leukemia subtypes.

R2.

Table 1 is set out in the 2016 ALL classification of the World Health Organization (WHO). Consequently, we consider it inappropriate to introduce modifications to the table.

Although Table 1 is quite informative for the subtypes, the review itself overly concentrates on cases of BCR-ABL1+ ALL or T-cell ALL.   The authors could discuss the other leukemia fusion gene subtypes mentioned – even only as potential candidates for SL treatment? 

For example, epigenetic SL has been proposed for MLL-r acute leukemias (mainly in AML), but PARP inhibitors, HDAC inhibitors etc may all be able to play a role in BCP-ALL treatment through SL and should be discussed in some more detail.

R3.

The manuscript focuses on BCR-ABL1+ ALL or T-cell ALL, in line with current synthetic lethality research that predominantly concentrates on these ALL subtypes. Table 2 exclusively lists confirmed cases of synthetic lethality. Considering that numerous synthetic lethal interactions are dependent on specific conditions and may not be universally conserved in all genetic backgrounds or cellular conditions, we are cautious in referring to reports of synthetic lethality in other leukemia types or cancer subtypes. However, we have incorporated a paragraph that addresses the point you mention and is presented in the manuscript as follows:

While current research predominantly emphasizes certain subtypes of ALL, it may be prudent to explore synthetic interactions within each of the subtypes outlined in the 2016 World Health Organization (WHO) ALL classification. This is particularly relevant for those subtypes that are associated with poor clinical prognosis for patients.

For Table 2, perhaps the title should also state "in ALL cell lines"?

R4.

As you suggested, it has been added to the title of the table.

In Table 2 it is quite difficult to interpret which of the genes shown to be involved in the different leukemia subtypes correspond to the observations detected after SL treatment.  Better spacing of the lines between the subtypes in the table may help?   Why is BCR-ABL1 fusion only shown for one Ph+ subtype?  Should the relevant TKI’s or other used inhibitors also be shown in the Table?

R5.

Following your suggestion, we enhanced the spacing between rows. Regarding your inquiries, the table exclusively displays genes identified as causing synthetic lethality based on the provided references. Consequently, the inclusion of the BCR-ABL1 gene is limited to Ph+ ALL, and details about the associated drugs are not included.

Minor:

Line 55: What is the percentage of overall relapse (what does high incidence mean here?).  Isn't leukemia relapse sub-type dependent?

R6.

The required information has been included and, although it is recognized that relapse is a factor depending on the subtype, as you mention, general data is used for practical reasons. In the manuscript it appears as follows:

Notably, the high incidence of relapses (In children it is around 10%, while in adults it is close to 50%) after initial treatments represents a major challenge, constituting the main cause of mortality in pediatric patients diagnosed with ALL.

Reviewer 2 Report

Please find my comments in the attached document

Comments for author File: Comments.pdf

Author Response

Dear Reviewer

We thank you for taking the time to review our work and for your valuable comments, which help us to improve its quality. Responses to their comments are given below, and all modifications to the manuscript are indicated in red text.

Reviewer 2

To authors

Lines 24 to 30. Do readers really need this basic information? I would suggest removing this

paragraph.

R1.

This section was added to facilitate understanding for people who are not experts in the field of leukemia, as highlighted by reviewer 1. Therefore, we consider it appropriate to keep this explanation.

Lines 32 to 35. Reference number 8 does not support this sentence, please check and consider removing or changing this reference.

R2.

This reference was removed.

Lines 36 to 37. Reference number 10 do not support this sentence, please check and consider removing this reference

R3.

This reference has been substituted with the article bearing doi: 10.1093/aje/kwaa215.

Lines 37 to 38. I cannot find how reference number 7 support this sentence, please check and, if appropriate, please specify where exactly in this excellent report we can read a persistent increase of B-ALL incidence

R4.

This reference has been substituted with the article bearing doi: 10.1093/aje/kwaa215.

Lines 55 to 57. It seems clear that reference number 15 does not fit here since you are talking about ALL and not AML.

R5.

This reference has been substituted with the article bearing doi: 10.1200/EDBK_280175, which explores treatments and relapses in ALL.

Lines 62 to 63. Would you consider adding an abbreviation for synthetic lethality (SL) or synthetic lethality interaction (SLI) to be used hereafter?

R6.

We prefer to keep the number of abbreviations to a minimum.

Lines 78 to 80. This sounds somehow redundant: “Third, synthetic lethal interactions are rare, requiring a large number of mutant gene pair combinations to be queried to be identified”

R7.

We believe this information is not redundant, but rather essential to elucidate the challenges in identifying synthetic lethal interactions and why such discoveries are inherently challenging.

Lines 97 to 99. Reference number 22 should not fit here since you are talking about ALL and not AML.

R8.

This reference has been substituted with the article bearing doi: 10.1158/2159-8290.CD-20-1503, which explores the therapeutic applications of synthetic lethality.

Lines 114 to 115. It would be appropriate to mention how the prognosis of this entity changed radically after the introduction of tyrosin kinase inhibitors

R9.

What you mentioned was added briefly and appears as follows:

Historically, this subgroup has shown inferior results compared to their Ph-negative (Ph-) counterparts, although this scenario has improved with the introduction of tyrosine kinase inhibitors [27].

Lines 118 to 122. Here you do not explain how Ph+ ALL is particularly susceptible to this combination (taselisib and olaparib) which seems to be related to genomic instability secondary to BCRA1 downregulation.

R10.

The sentence was completed as follows:

Synthetic lethality has been reported in Ph+ B-ALL cell lines when combining a phosphoinositide-3-kinase (PI3K) inhibitor, such as taselisib, with olaparib, a poly(ADP-ribose) polymerase (PARP) inhibitor. This is because taselisib causes a reduction in breast cancer 1 (BRCA1) and breast cancer 2 (BRCA2) levels that affects homologous recombination (HR) repair and olaparib inhibits base excision repair (BER). Both DNA repair pathways play a crucial role in preserving genomic stability. Thus, Ph+ ALL cells show increased vulnerability to genetic instability secondary to BCRA1 downregulation and BER inhibition, ultimately leading to apoptosis.

Lines 129 to 138. One comment similar to the previous one: here, the “common reader” would ask for some reason to explain how Ph+ B-ALL is particularly sensitive to this combination (nilotinib plus MKC-8866) which seems to be related to its specific IREα-related survival dependency

R11.

The sentence was completed as follows:

In an effort to improve the efficacy of Ph+ B-ALL treatment with a different approach, a study analyzed the combination of BCR-ABL1 inhibition using the tyrosine kinase in-hibitor (TKI) nilotinib, together with the IRE1α RNase domain inhibitor MKC-8866. IRE1α assumes a critical role in the unfolded protein response (UPR) by performing messenger RNA splicing of the transcription factor XBP1. XBP1 targets encode proteins that contribute to improved folding and quality control of endoplasmic reticulum (ER) proteins. This therapy fights the disease through specific and complementary mechanisms of action affecting UPR proteins, multiple inducers of apoptosis and negative regulators of the cell cycle.

Lines 152 to 157. Should you consider explaining that both WEE1 and CHK1/CHK2 are cell cycle checkpoint kinases and how their inhibition induces cell death? The reader would find this more amenable

R12.

Based on your comment, this information was added and appears as follows in the manuscript:

Particularly in B-ALL cells with elevated WEE1 levels, one study mentioned synthetic lethality resulting from the combination of the CHK1/CHK2 inhibitor PF-0477736 and the WEE1 inhibitor adavosertib (AZD1775) [32]. Both CHK1/2 and WEE1 are cell cycle checkpoint kinases, and their sustained inhibition leads to cell death [31]. Tus, the ra-tionale for this drug regimen was to activate the S-phase checkpoint and induce DNA damage, thereby elevating replicative stress and causing the collapse of the replicative forks [32].

Lines 157 to 160. Here you should stress how TCF3-HLF ALL represents a very aggressive ALL subtype with a very poor outcome with current standard treatment approaches, including chemotherapy a stem cell transplantation

R13.

What you mention has been added and appears as follows in the manuscript:

Notably, TCF3-HLF ALL represents an exceptionally aggressive subtype characterized by a remarkably poor outcome with existing standard treatment modalities, including chemotherapy and stem cell transplantation [33].

Line 199; Table 2. You should find a way to separate related information; in the present format it is difficult to relate some genes and observations to their corresponding reference. Please try to clearly separate each line within the table

R14.

Table 2 has been improved with wider spacing between rows to make it easier to read and understand.

Lines 212 to 214. I would suggest: The underlying cause of this effect can be attributed, at least in part, to reduced dihydrofolate reductase (DHFR) expression caused by sirolimus-induced decreased cyclin D1 levels

R15.

Based on your comment, the sentence was modified.

Lines 230 to 232. Consider removing “was” from the following sentence: “Other potential targets in Ph+ ALL that could be considered as potential synthetic lethality was predicted along with ABL1 hyperactivity in a CRISPR-based genotoxic screen against DNA damaging agents are…”

R16.

Based on your comment, the sentence was modified as follows:

Other potential targets in Ph+ ALL, which could be explored for their potential synthetic lethality in conjunction with ABL1 hyperactivity, were identified using a CRISPR-based genotoxic assay against DNA-damaging agents.Among these targets are the fu-cose-1-phosphate guanylyltransferase (FPGT)-cardiac troponin I-interacting kinase (TNNI3K), NADH:ubiquinone oxidoreductase subunit C1 (NDUFC1), β-1,3-galactosyltransferase 6 (B3GALT6), neuromedin U receptor 2 (NMUR2), AP2 as-sociated kinase 1 (AAK1), zinc finger X-linked duplicated A (ZXDA), calpastatin (CAST), γ-aminobutyric acid type A receptor subunit γ2 (GABRG2), GFB-induced factor 2 (TGIF2), crystallin γS (CRYGS) and family with sequence similarity 105, member B (FAM105B)

Lines 254 and 264. Please, consider KMT2A-rearranged instead of MLL-rearranged or use both terms

R17.

Following your suggestion, we have replaced "MLL-rearranged" with "KMT2A-rearranged".

Round 2

Reviewer 1 Report

the paper is now ready for publication

Reviewer 2 Report

No additional comments