Small nucleolar RNAs in plasma extracellular vesicles and their discriminatory power as diagnostic biomarkers of Alzheimer's disease

May combine two claims. The quote only partly states this claim.

The AUC values for SNORD115 (0.861), SNORD116 (0.886), and combined (0.947) exceeded the threshold of 0.8.

“The values of AUC for SNORD115–0.861, as well as for SNORD116–0.886 exceeded the threshold, thus according to widely accepted classification within the range of “outstanding” (Konstantinou et al., 2012; Mandrekar, 2010).”

• Also stated in: § Verification of differentially enriched SNORDs in plasma EV of an independent group of AD patients and NC · plain

  “The value of combined AUC calculated for both, SNORD115 and SNORD116, was 0.947 (Fig. 3E–F).”

• cites Konstantinou et al., 2012 sourceDisputed

  The Methods (‘Performance of the diagnostic model’) classify AUC values from 0.8 to below 0.9 as ‘Excellent discrimination,’ not ‘Outstanding discrimination.’ The cited paper supports that 0.861 and 0.886 exceed 0.8, but contradicts the citing sentence’s classification of those values as outstanding. It does not report the SNORD results. (LLM, conf=0.90)

  Checked by a reasoning model — may be wrong.

  Experimental — accuracy not yet measured
• cites Mandrekar, 2010 sourceNot checked — no full text of the source

May combine two claims.

There has been only one study reporting a combined AUC of 0.98 for p-tau and Aβ42/40 based on CSF ELISA assays, plus a signature of combined 3 miRNAs and 3 piRNAs from CSF deep sequencing.

“Moreover, there has been only one study reporting a combined AUC of 0.98 for p-tau and Aβ42/40 with calculations based on CSF ELISA assays, plus a signature of combined 3 miRNAs +3 piRNAs with scores derived by deep sequencing of RNA isolated from CSF samples (Jain et al., 2019).”

• cites Jain et al., 2019 sourceSupported

Probably not a substantial claim: may not pass S3 (load-bearing).

The plasma piRNAs identified in this study did not overlap with those in Jain et al.'s study of EV isolated from CSF.

“The fact that the identified plasma piRNAs in our study didn’t overlap with Jain et al. study performed on EV isolated from CSF (Jain et al., 2019) is not surprising.”

• cites Jain et al., 2019 sourceSupported

Probably not a substantial claim: may not pass S3 (load-bearing).

Engineered expression of members of SNORD115 in brain tissues other than brain parenchyma (choroid plexus) has been reported.

“Engineered expression of members of SNORD115 in brain tissues other than brain parenchyma – choroid plexus, has also been reported (Raabe et al., 2019).”

• cites Raabe et al., 2019 sourceSupported

May combine two claims.

SNORD115 and SNORD116 are clustered in intronic regions of an imprinted host gene and are known to be expressed predominantly (SNORD116) or exclusively (SNORD115) in brain.

“SNORD115 and SNORD116 are clustered in intronic regions of an imprinted host gene and are known to be expressed predominantly – SNORD116, or exclusively – SNORD115, in brain (Cavaillé, 2017; Chamberlain, 2013; Chamberlain and Lalande, 2010a; Chung et al., 2020; Driedonks et al., 2018; Irimie et al., 2018; Lässer et al., 2016).”

• cites Cavaillé, 2017 sourceNot checked — no full text of the source
• cites Chamberlain, 2013 sourcePartly supported

  The Non-coding RNAs section identifies SNORD115 and SNORD116 as clusters processed from introns of the SNURF-SNRPN host transcript. It says the portion hosting SNORD115 is expressed only in brain. The Human-Mouse shared synteny section says human non-coding RNAs in the region before IPW are expressed in a variety of cell types; it does not establish that SNORD116 expression is predominantly in brain. Page numbers are not available in our copy. (LLM, conf=0.85)

  Experimental — accuracy not yet measured
• cites Chamberlain and Lalande, 2010aNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Probably not a substantial claim: may not pass S3 (load-bearing).

The authors have not been able to find a report that SNORD115 or individual members of the SNORD115 cluster were released in plasma EV of patients with lung cancer.

“However, we have not been able to find a report that SNORD115 or individual members of SNORD115 cluster were released in plasma EV of patients with lung cancer (Nossent et al., 2019; Steinbusch et al., 2017).”

• cites Nossent et al., 2019 sourceCould not verify

  The Results (section 2.1) and Methods (section 4.7; page numbers unavailable) concern plasma SNORD112, SNORD113.2, SNORD113.6, and SNORD114.1 in patients with peripheral artery disease. Searches for SNORD115, extracellular vesicles, and lung cancer found no passage addressing whether a report of SNORD115 release in plasma EV exists. (LLM, conf=0.76)

  Experimental — accuracy not yet measured
• cites Steinbusch et al., 2017 sourceCould not verify

  The Results and Discussion address serum snoRNAs in joint ageing and osteoarthritis, particularly SNORD116, but do not address SNORD115 in plasma extracellular vesicles from patients with lung cancer. Searches for SNORD115 and lung-cancer plasma extracellular vesicles found no passage in our copy establishing the citing authors’ statement about the absence of a report. (LLM, conf=0.75)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing).

We have recently reported a correlation of snoRNAs with APOE genotype in postmortem brain tissue of AD patients.

“We have recently reported a correlation of snoRNAs with APOE genotype in postmortem brain tissue of AD patients (Fig. 4) (Lefterov et al., 2019).”

• cites Lefterov et al., 2019 sourceSupported

May combine two claims.

SNORD115 is expressed exclusively in brain and specifically in neurons.

“this cluster of ncRNAs is expressed exclusively in brain and specifically in neurons (Bortolin-Cavaille and Cavaille, 2012; Cavaillé, 2017; Chamberlain, 2013; Chamberlain and Lalande, 2010b; Chung et al., 2020; Keshavarz et al., 2020; Wevrick et al., 1994).”

• cites Bortolin-Cavaille and Cavaille, 2012 sourcePartly supported

  In Results and Discussion (Figure 2; page unavailable), the paper detects SNORD115 in brain RNA and uses liver as a negative control. It does not establish expression exclusively in the brain. The Conclusion only speculates that production of SNORD115 derivatives may be restricted to neuronal tissues; it does not show that SNORD115 itself is expressed specifically in neurons. (LLM, conf=0.83)

  Experimental — accuracy not yet measured
• cites Cavaillé, 2017 sourceNot checked — no full text of the source
• cites Chamberlain, 2013 sourcePartly supported

  The Non-coding RNAs section (page not available) says the brain-only portion of the SNURF-SNRPN transcript serves as the host for SNORD115/HBII-52, supporting brain-specific expression. It does not say that SNORD115 expression is restricted to neurons within the brain; the Conclusions instead call for comparisons of neuronal and non-neuronal RNAs. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing).

CSF biomarkers' clinical utility is considered appropriate only in select clinical situations.

“currently, their clinical utility is considered appropriate only in select clinical situations – in the setting of a dementia clinic, in patients with early-onset, progressive or unexplained MCI or with comorbidities that add uncertainty to the diagnosis of AD (Shaw et al., 2018).”

• cites Shaw et al., 2018 sourcePartly supported

  Section 8 (Implementation of these AUC; page unavailable) recommends testing by dementia experts and identifies progressive, unexplained MCI, early-onset MCI or dementia, and possible AD with diagnostic uncertainty from comorbidities as appropriate situations. But it also identifies three other appropriate indications, so the sentence's restrictive list is incomplete; the paper specifies dementia experts, not necessarily a dementia-clinic setting. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing).

There is evidence that malignant cells of lung cancer tissue in primary cultures and lung cancer cell lines express members of the SNORD115 cluster, in addition to many other SNORDs and SNORAs.

“Although it has been reiterated in recent publications, (Cavaillé, 2017; Chung et al., 2020; Deogharia and Majumder, 2018), there is evidence that malignant cells of lung cancer tissue in primary cultures and lung cancer cell lines express members of SNORD115 cluster, in addition to many other SNORDs and SNORAs (Gao et al., 2014; Liao et al., 2010; Mourksi et al., 2020).”

• cites Gao et al., 2014 sourcePartly supported

  Table 3 (p. E628) identifies SNORD115-32 and SNORD115-41 alongside other SNORDs and SNORAs in lung tumor tissue. The Patients and clinical specimens section describes surgically removed and frozen tissues, not primary cultures or lung cancer cell lines; the paper also does not establish expression specifically in isolated malignant cells. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Liao et al., 2010 sourcePartly supported

  The Results section reports expression of several SNORDs and SNORA42 in resected NSCLC tissues. It does not report SNORD115-cluster expression or experiments on primary cultures or lung cancer cell lines; the Methods section describes surgical tissue specimens and plasma samples instead. Page numbers are unavailable in our copy. (LLM, conf=0.89)

  Experimental — accuracy not yet measured
• cites Mourksi et al., 2020 sourcePartly supported

  Sections 4.1 and 5 report SNORA and SNORD expression in lung cancer tissue and cell lines. But section 2.3 mentions SNORD115 only as part of a locus associated with Prader–Willi syndrome; the cited passages do not establish SNORD115-cluster expression in lung cancer cells. Section 4.1 describes primary surgical specimens, not primary cultures. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

May combine two claims.

In meningioma samples, members of the SNORD115 cluster have not been identified as up-regulated, supporting their exclusive neuronal origin.

“Interestingly, in meningioma samples – members of SNORD115 cluster have not been identified as up-regulated, supporting their exclusive neuronal origin (Jha et al., 2015; Rynkeviciene et al., 2018).”

• cites Jha et al., 2015 sourceNot checked — no full text of the source
• cites Rynkeviciene et al., 2018 sourceCould not verify

  The Discussion (page unavailable) describes snoRNA expression in pediatric gliomas, not meningioma samples. Searches for SNORD115, meningioma up-regulation, and neuronal origin found no passage addressing the attributed claim in our copy. (LLM, conf=0.73)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Blood tests to measure Aβ peptides, total and phospho-tau have the potential for screening asymptomatic individuals and aiding diagnosis of symptomatic patients.

“Blood test to measure Aβ peptides, total and phospho-tau are much more appropriate tools, and regardless of their own and significant challenges, they have the potential for screening asymptomatic individuals and to aid in diagnosing symptomatic patients (Ashton et al., 2021).”

• cites Ashton et al., 2021 sourcePartly supported

  The Discussion (page not supplied) says plasma p-tau could aid diagnosis in patients with dementia and that plasma Aβ may help predict preclinical AD. But the Results, Phase 1 (page not supplied), explicitly says blood total tau is not diagnostically useful for AD, so the claim overextends that potential to total tau. (LLM, conf=0.90)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Differential expression of snoRNAs has been observed in brain of AD model mice and was associated with initial progression of amyloid depositions.

“Differential expression of snoRNAs has been observed in brain of AD model mice and were associated with the initial progression of amyloid depositions (Gstir et al., 2014).”

• cites Gstir et al., 2014 sourcePartly supported

  The Results and Discussion section on young 3xTG mice reports differential expression of snoRNAs e307 and e470 in cortical samples. It says these changes occurred before β-amyloid plaque formation and were no longer observed after plaques formed; it does not establish an association with the initial progression of amyloid deposition. No page number is available in our copy. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing).

Members of SNORD115 and SNORD116 clusters have been reported upregulated in patients with multiple myeloma, chronic lymphocytic leukemia, or acute lymphoblastic leukemia.

“Members of SNORD115 and SNORD116 clusters have been also reported upregulated in patients with multiple myeloma, chronic lymphocytic leukemia (Ronchetti et al., 2013; Ronchetti et al., 2012), or acute lymphoblastic leukemia (Vendramini et al., 2014).”

• cites Ronchetti et al., 2013 sourcePartly supported

  The Results section on high-risk CLL reports higher SNORD116-18 expression in a subgroup of CLL patients, compared with a lower-expression subgroup. It does not establish that SNORD116 members are generally upregulated in CLL patients relative to healthy cells, and it does not report SNORD115 upregulation in CLL. The multiple-myeloma and acute-lymphoblastic-leukemia portions are not attributed specifically to this paper. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites Ronchetti et al., 2012 sourceSupported
• cites Vendramini et al., 2014Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

May combine two claims.

For both SNORD115 and SNORD116, there was a significant difference between AD and control samples in ddPCR absolute quantification.

“As shown on Fig. 3 A and C, for both SNORD115 and SNORD116 there was a significant difference between AD and control samples.”

May combine two claims.

In postmortem brain samples, 102 SNORD genes were identified, and 12 SNORD transcripts were differentially expressed between AD-E4+ and AD-E3/3.

“Here, we compared transcriptomic profiles of APOE3/3 vs APOE4+ (see demographics on Table 3) and identified 102 SNORD genes (Fig. 4A, Supplementary Table 2).”

• Also stated in: § Differentially enriched SNORDs in brain of AD patients and NC · plain

  “There were 12 SNORD transcripts differentially expressed between AD-E4+ and AD-E3/3 (Fig. 4A–B).”

May combine two claims.

For both SNORD115-32 and SNORD116-29, the calculated AUCs were poor and therefore they did not justify further testing.

“Importantly, for both SNORDs we calculated poor AUCs and therefore they did not justify further testing.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Because APOE genotype could be a factor, HDL:RNA complexes likely have an impact on the final recovery of ncRNAs processed for sequencing.

“Because the APOE genotype could be a factor, HDL:RNA complexes likely have an impact on the final recovery of ncRNAs processed for sequencing.”

Probably not a substantial claim: may not pass S3 (load-bearing).

The 39 samples were randomly chosen from a series collected routinely at ADRC Pittsburgh between 2015 and 2018.

“All 39 samples – APOE3/3 and APOE 4/+ genotypes, were randomly chosen from a series collected routinely at ADRC Pittsburgh between 2015 and 2018 (Table 1).”

Probably not a substantial claim: may not pass S3 (load-bearing).

RNA concentrations did not differ between plasma EVs from AD patients and NC.

“The concentration of RNA was measured within the range of 41.6–141 pg/μl (mean 73.2 pg/μl) with no difference between RNA concentrations of plasma EVs from AD patients and NC (unpaired two-sample t-test; GraphPad Prism v8.4.2).”

May combine two claims.

The enrichment of EV obtained by ultracentrifugation was confirmed by detecting four EV markers (Flotillin, HSP70, CD63, TSG101) and the lack of BiP.

“The enrichment of EV obtained by ultracentrifugation was confirmed by detecting four different EV markers by Western blot: Flotillin, heat shock protein 70 (HSP70), the CD63 antigen and tumor susceptibility gene 101 (TSG101, a component of the ESCRT-I complex, a vesicular trafficking process regulator) (Fig. 1D) and the lack of BiP (Endoplasmic reticulum chaperone BiP).”

Probably not a substantial claim: may not pass S3 (load-bearing).

Members of SNORD113 and SNORD114 clusters identified as differentially enriched are relatively highly expressed in liver, breast, ovary and prostate.

“the members of SNORD113 and SNORD114 clusters identified differentially enriched in our study are relatively highly expressed in liver, breast, ovary and prostate (Dsouza et al., 2021).”

• cites Dsouza et al., 2021 sourceNot checked — no full text of the source

May combine two claims.

No difference in EV size or concentration was noticed between AD and NC groups.

“No difference of EV size or concentration was noticed between AD and NC groups (Fig. 1B–C).”

May combine two claims.

The clinical diagnosis of dementia used by the University of Pittsburgh ADRC has a 98% sensitivity and 88% specificity for AD.

“it has shown a 98% sensitivity and 88% specificity for AD (Lopez et al., 2000a; Lopez et al., 2000b).”

• cites Lopez et al., 2000aNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Lopez et al., 2000bNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Each ddPCR assay included a negative control for each RT reaction and a no-template control.

“Each assay included a negative control for each RT reaction and a no-template control according to the protocol provided by BioRad.”

May combine two claims.

The results presented here point to a newly discovered association of APOE genotype, increased transcriptional activity of imprinted control regions, facilitated release of SNORDs in plasma EV and AD pathogenesis.

“Although, the outcomes of much larger independent validation studies are difficult to predict, the results presented here point, at least, to a newly discovered association of APOE genotype, increased transcriptional activity of imprinted control regions, facilitated release of SNORDs in plasma EV and AD pathogenesis.”

• AssumptionThe authors state this as an assumption: Members of SNORD115 in EV of plasma samples presented in this study come from brain.

Probably not a substantial claim: may not pass S3 (load-bearing).

The study was approved by the Institutional Review Board of the University of Pittsburgh.

“The study was approved by the Institutional Review Board of the University of Pittsburgh.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

The differential expression of SNORD115 and SNORD116 genes in postmortem brains of AD patients, even without comparison to non-demented controls, is a clear indication that a genotype effect is likely to exist.

“The differential expression of SNORD115 and SNORD116 genes in postmortem brains of AD patients, however, even though without comparing the levels to postmortem brains of NC (Fig. 4) is a clear indication that a genotype effect is likely to exist.”

Classified as probably not a substantial claim.

Current criteria for AD diagnosis require well-defined clinical phenotype and pathophysiological biomarkers consistent with AD pathology.

“Current criteria for AD diagnosis require well-defined clinical phenotype and pathophysiological biomarkers consistent with AD pathology (Dubois et al., 2014).”

• cites Dubois et al., 2014 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing).

The Nanosight instrument was calibrated before each batch of samples using 100 nm latex beads.

“The instrument was calibrated before each batch of samples using 2 μL of 100 nm latex beads diluted in 1 ml of PBS.”

May combine two claims.

The EV isolation procedure used (differential ultracentrifugation with initial low speed centrifugation and filtering) provides intermediate recovery and specificity of EV, so contamination of the pellet with vesicles smaller than 30 nm or larger than 100 nm cannot be excluded.

“Because the differential ultracentrifugation with initial low speed centrifugation and filtering provides intermediate recovery and specificity of EV (Théry et al., 2018), we cannot exclude the small possibility of contamination of the pellet with vesicles smaller than 30 nm, neither with those larger than 100 nm.”

• cites Théry et al., 2018Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.