Case Report of Small Cell Carcinoma of the Ovary, Hypercalcemic Type (Ovarian Rhabdoid Tumor) with SMARCB1 Mutation: A Literature Review of a Rare and Aggressive Condition

Round 1

Reviewer 1 Report

I was happy to see that the Authors were able to re-submit a revised version of this interesting case report. I would like to thank the Authors for thoroughly addressing my previous comments and for comprehensively answering my questions. In my opinion, the revised version of the manuscript reads much better, and the Discussion section in particular appears to be more clear and precise. 

Because the manuscript underwent modifications and several new elements were added, I have four additional comments that I would like the Authors to address prior to final acceptance and publication. These comments are minor and should hopefully be easy to address. They are listed below:

1) Page 3 - line 59: a new piece of information has been added: "cytokeratin staining was positive in areas where INI-1 expression was preserved". This is unfortunately quite confusing, as the existence of tumor areas with preserved INI-1 expression is not discussed or explained elsewhere in the text (and I believe it was not mentioned in the initial version of the manuscript). It could raise additional questions (e.g., is this really a SCCOHT and not a dedifferentiated component of another tumor? Is it a SCCOHT arising in a germ cell tumor?). It is also unusual for SCCOHT or for rhabdoid tumors not to show a complete loss of the deficient SWI/SNF protein. 

At this point, I would suggest two possible ways of moving forward: a) the Authors could describe this intriguing finding in more detail (e.g. describe what percentage of tumor cells showed retained INI-1 expression, and whether these areas were different in any way from the rest of the tumor); or b) if this was a very minor phenomenon and/or only seen on the initial biopsy, the Authors could remove the statement "in areas where INI-1 expression was preserved" and, when describing the cytokeratin staining, simply use the term "...focal cytokeratin expression" without going into further detail.

Also, please note that the legend of Figure 5 requires a correction (should read "IHC" instead of "H&E").

2) Figure 6A: the microphotograph appears to be out of focus, and it is hard to appreciate whether the staining is nuclear. Could the Authors please provide a better quality image and at a slightly higher magnification? In addition, the figure legend requires some corrections ("SMARCA4" instead of "SMARC4" and "IHC" instead of "H&E").

3) Figure 6B: based on this microphotograph, in my opinion, SMARCA2 expression (expected to be nuclear) actually seems to be negative in tumor cells (as compared to tumor infiltrating lymphocytes for example). However, the background and cytoplasmic "noise" make it quite difficult to evaluate this IHC. I would suggest two possible ways of moving forward: a) the Authors could show a slightly higher magnification (to better demonstrate the negative tumor nuclei) and state in the text that SMARCA2 expression was negative in tumor cells; or b) if the Authors do not agree with this interpretation of the staining, then I would suggest removing Figure 6B and removing the SMARCA2 result altogether from the text (line 78). 

If the Authors decide to keep Figure 6B as part of the manuscript, the figure legend would require some corrections ("SMARCA2" instead of "SMARC2", "lack of nuclear SMARCA2 expression" instead of "expression retained" and "IHC" instead of "H&E"). 

4) In the response letter, the Authors agreed that dedifferentiated carcinoma of the ovary is a possible differential diagnosis to consider. However, I fully  agree with the Authors that many elements (patient's age, the absence of a well-differentiated component, the presence of follicle-like structures in the tumor, and the low tumor mutation burden) favor the diagnosis of SCCOHT over dedifferentiated carcinoma. I would nevertheless suggest to include one sentence summarizing these exact considerations in the manuscript, for example in the Case Report section (e.g. line 151). 

As mentioned above, once these last points have been addressed, I believe this interesting manuscript will be perfectly suitable for publication in Current Oncology.

Author Response

Please see the attachment.

 

It was also suggested to include, in the text, the approval for publication from the patient. Find attached this information.

 

Best regards

Author Response File: Author Response.docx

This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.

Round 1

Reviewer 1 Report

Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) is an aggressive malignancy that occurs in young women, is characterized by recurrent loss-of-function mutations in the SMARCA4 gene, and for which effective treatments options are lacking. 
Occasional tumors exhibit loss of SMARCB1 instead of SMARCA4. Mutation of the SMARCB1 gene occurs frequently in soft tissue sarcomas
(synovial sarcomas, rhabdoid tumors), and in a small portion of SCCOHT. Thus I suggest to change the tittle by adding phrase ovarian rhabdoid tumor:
Case Report of Small Cell Carcinoma of The Ovary, Hypercalcemic Type (ovarian rhabdoid tumor): A Literature Review of a Rare and Aggressive Condition 

Reviewer 2 Report

The case report by Maria Fernanda Evangelista Simoes and coworkers describes a 19 year old patient diagnosed with small cell carcinoma of the ovary of the hypercalcemic type (SCCOHT). Importantly, instead of a loss-of-function SMARCA4 alteration which is classically found in these tumors, this patient’s tumor harbored a biallelic truncating SMARCB1 mutation. The authors report comprehensive clinical, radiologic, pathologic and genomic findings in this patient, as well as the details of the clinical management. 

Given the unusual genomic findings in this tumor, and the rarity of SCCOHT overall, this case is indeed important to report to the medical and scientific community. The manuscript could be appropriate for publication in Current Oncology, pending addressing the below specific comments. In addition, the Discussion section needs to be improved overall.

1) The presence of a somatic SMARCB1 alteration instead of the expected SMARCA4 alteration in a SCCOHT is in my opinion the key finding in this study. As such, the SMARCB1 mutation should appear in the title, and should also be clearly mentioned in the abstract.

In addition, I would suggest to mention the significance of this finding for routine practice in the Discussion or the Conclusion. For example, the authors could emphasize the importance of performing INI1/SMARCB1 IHC on undifferentiated ovarian tumors in young patients that show retained SMARCA4 expression.

2) As rightly pointed out by the authors in the Discussion, SCCOHT are typically characterized by a low tumor mutation burden (TMB) and a remarkably stable genome. Given the unique nature of this case, it would be important to provide an assessment of these two parameters. A low TMB and a stable genome would further support the diagnosis of SCCOHT. Conversely, the opposite findings would be unusual for SCCOHT and perhaps warrant additional discussion. Given that DNA sequencing of the tumor was performed using a 409-gene panel, it should technically be possible to obtain an estimate of the genomic copy number profile and the TMB. I strongly encourage the authors to try to include this information.

3) An obvious question raised by this unusual case is, how confident can we be that this tumor is indeed a SCCOHT? Although the pathology aspects and clinical presentation appear very consistent with SCCOHT, some additional clarifications would be helpful:

- ER and PR are probably expected to be negative in SCCOHT (PMID: 32156746). Could the authors better describe the degree of positivity observed in this case, and perhaps discuss based on the literature whether they think this is an issue?

- Could the authors describe in more detail the pattern of cytokeratin staining?

- Did the authors observe any cells with a rhabdoid morphology?

- Could the authors confirm that no tumor was present in the endometrium?

- Could the authors discuss whether another entity such as dedifferentiated carcinoma of the ovary (PMID 16306785) should be considered as a differential diagnosis, or whether this is unlikely?

4) Dual loss of expression of SMARCA4 (BRG1) and SMARCA2 (BRM) by IHC is a diagnostic tool for SCCOHT. The authors state that SMARCA4 (BRG1) expression was retained. I would suggest to include a picture of this IHC in addition to INI1/SMARCB1, to formally demonstrate the strong retained expression. If possible, I would also encourage the authors to perform SMARCA2 (BRM) IHC on this tumor. Was SMARCA2 (BRM) expression completely negative, as classically seen in SCCOHT and in some SMARCB1-deficient rhabdoid tumors, or was it heterogeneous, as can be seen in a subset of SMARCB1-deficient rhabdoid tumors?

5) Some information provided in the Discussion seems to be either incorrect or incomplete. In particular, please reconsider the following points:

a) Stating that SMARCB1 mutations have previously been reported in rare cases of SCCOHT does not give a clear idea of the number of such cases (page 8, line 180; page 8, lines 213 and 215). To the best of my knowledge, three cases of SMARCB1-deficient SCCOHT have been reported (Karnezis et al. PMID: 26356327). If the authors are aware of other published cases of SMARCB1-deficient SCCOHT, then these papers should be explicitly cited and the cases “counted”. 

b) To my knowledge, the statement that SMARCA2 mutations have been identifed in SCCOHT (page 8, line 181) is not correct. SMARCA2 “deficiency” described in SCCOHT is due to the lack of protein expression, but not to deleterious mutations of the gene. If this statement is incorrect, the authors should remove it. If the authors are aware of publications specifically reporting SMARCA2 mutations in SCCOHT, then these should be cited. The reference [12] cited page 8, line 181 does not report SMARCB1 or SMARCA2 mutations in SCCOHT.

c) Line 215, the authors state that SMARCB1 mutations can occur in synovial sarcoma. This is technically incorrect: synovial sarcoma shows decreased expression of INI1 (SMARCB1) at the protein level, which can be evidenced by IHC, but this is not due to SMARCB1

d) Line 215, the list of malignancies which can display SMARCB1 mutations is very short. I would suggest to specifically emphasize rhabdoid tumors (please also see point 6 below), and perhaps to list some other tumors that display recurrent SMARCB1 alterations (such as medullary carcinoma of the kidney, epithelioid sarcoma, undifferentiated chordoma, epithelioid MPNST, myoepithelial carcinoma...) – and of course the gynecologic malignancies that the authors rightly mention line 217.

e) Line 150, “SCCOHT is frequently diagnosed at an early stage”. Is this really the case? Most papers state that these tumors are diagnosed at an advanced stage. Please cite specific references to support this “early stage” statement. If this is not supported by the literature, please remove.

f) The statement that the approach to SCCOHT is “based on case reports” (page 8, line 224) sounds odd given that prospective clinical trials are mentioned later in the same section of the Discussion.

h) Line 237: “an improvement in grades 3/4 adverse events”: the cited study actually appears to say the opposite, i.e. that grade 3/4 adverse events were frequent with the dose-intensive regimen (even though they were manageable).

6) Overall, although the Discussion encompasses important concepts relative to SCCOHT, it could benefit from being re-written or re-structured for more clarity. For example:

a) In paragraphs 4 through 8 of the Discussion, the text “skips” back and forth between similar concepts, and some concepts are repeated several times.

b) The literature on genomic and molecular aspects of SCCOHT is not clearly presented. I would suggest to dedicate a separate paragraph to this very important topic (perhaps right after the clinical and pathology findings), instead of “abruptly” introducing it at the end of a pathology paragraph.

The identification of recurrent, biallelic, deleterious SMARCA4 alterations in nearly all cases of SCCOHT was a breakthrough finding for the understanding of these tumors, and should be clearly presented. The four seminal papers which first identified these SMARCA4 alterations in SCCOHT (PMID 24375037, 24658002, 24658001, 24658004) should be adequately cited. The importance and high prevalence of germline alterations could also be mentioned here. Next, the authors could be emphasize that SMARCA4 alterations lead to a loss of expression of the SMARCA4 (BRG1) protein, and that this is also accompanied by the loss of expression of the SMARCA2 (BRM) paralog (original paper: 26356327), albeit without underlying SMARCA2 mutations. Next, the low TMB and genomic stability can be discussed to complete this “molecular” paragraph (Lin et al., Auguste et al.). Functional studies and any additional points can be mentioned afterwards.

c) Page 8 line 190, the sentence “There is a strong correlation between SCCOHT and germline and somatic deleterious mutations in SMARCA4” is imprecise. The authors should mention the exact prevalence of germline SMARCA4 alterations in SCCOHT found in the literature (which is around 40%) and cite the appropriate papers.  

d) Page 8, line 212: the sentence “Over 95% of these neoplasms exhibit loss of nuclear immunoreactivity, making this marker an important diagnostic tool” is not clear, as it comes right after a sentence mentioning SMARCB1. It sounds like “95% of SCCOHT exhibit loss of nuclear immunoreactivity for SMARCB1”, which is obviously not what the authors meant here.

e) Page 8 and 9, a clinical trial published in 2020 is being described before the one published in 2007. Please consider describing these trials in chronological order.

6) This interesting case begs the question about similarities and differences between SCCOHT and malignant rhabdoid tumors (RTs). RTs are the bona fide model of tumors driven by SMARCB1 inactivation, which is important to emphasize (in the current version of the manuscript, rhabdoid tumors are only mentioned once, very briefly). Some authors have suggested that SCCOHT and RTs should be considered as a unique entity (PMID 24752781). Conversely, a recent study (PMID 33999421) provided molecular arguments for their continuous separate classification, and also showed that SMARCA4-deficient RTs (a rare subset) show intermediate molecular features between SMARCB1-deficient RTs and SMARCA4-deficient SCCOHT. It would be intriguing to speculate where this case of a SMARCB1-deficient SCCOHT would "fall" on this spectrum of tumors.

I would suggest that the authors mention these considerations in the Discussion and reference the appropriate papers.

Minor comments:

1) Line 179: “the histogenesis of this tumor remains uncertain” – it could be useful to mention here the hypothesis of a germ cell origin and possible association with minute foci of germ cell tumor (PMID 24375037, 28130795)

2) Line 184: “The dual deficiency in core members of the SWI/SNF complex (e.g., SMARCA4 and SMARCA2 or SMARCB1 and SMARCA2) can induce dedifferentiation from a normal cell or a low-grade tumor into an aggressive high-grade tumor with small cell and/or rhabdoid features”. This sentence, used in the paper by Karnezis et al. (PMID 26356327), is hypothetical. Please consider rewording (e.g. “it has been hypothesized that....”) Also, technically, only SMARCB1 is a “core” member; SMARCA4 and SMARCA2 are catalytic subunits. Perhaps saying “key” members would be safer.

3) Page 9, line 249: although the trial NCT02601950 did show clinical activity in 2 patients with SCCOHT, stage 2 futility was not passed. Please consider mentioning that only a subset of patients with SCCOHT is likely to benefit from EZH2inh.

4) Line 57: “Focal positivity of calretinin was also observed, favoring the hypothesis of epithelial neoplasia of gynecological origin. Thus, it was not possible to conclude primary tumor origin based on morphological and Immunohistochemistry analysis of the tumor sample taken by incisional biopsy.” – these two sentences appear contradictory, please reconsider.

5) Line 175: “The immunohistochemistry of SCCOHT reveals positivity...” – I would suggest to call this variable positivity.

6) Minor wording or typing issues:

- line 41: “lymph node” (nodes?)

- line 52: “fusiform” (spindle shaped may be more commonly used?)

- line 61: “was then submitted” (is “submitted” the correct word here?)

- line 67: “The surgical specimen had an ovary...” (consider rewording “had”)

- line 69: “Follicle-like spaces with eosinophilic secretions were seen.” (consider using a transition between gross and microscopic description, e.g. “On microscopic examination, ...”)  

- line 73: “However, it maintained a preserved expression for SMARCA4” (consider rewording, e.g. “showed retained expression of SMARCA4”)

- line 90: “sexual cords” (sex cords?)

- line 100: “Then there was...” (please consider a different beginning to this sentence)

- line 187: “Dual loss of SMARCA4 and SMARCA2 can happen in SCCOHT...” (“can happen” does not seem appropriate – this dual loss is a defining, recurrent feature)

- line 194: “The search of SMARCA4 is realized...” – wording is not clear. Please consider rewording (e.g. “the search for SMARCA4 alterations”) and using a different word (“performed”?) instead of “realized”

- line 198: I believe this study is not a “clinical trial”

- line 205: “diagnosis is reconsidered” (diagnosis be considered)

- line 206: “along with sequencing of other related genes as a SMARCB1 mutation that has been reported in a case of SCCOHT” (please consider rephrasing)

- Line 254: "adapted" (suitable?)

- line 255: “Moreover, more clinical trials are needed...” – “moreover” does not seem to be the right word here

- line 266: “had disease progression” – please consider a different word than “had"