Modification of antigen-encoding RNA increases stability, translational efficacy, and T-cell stimulatory capacity of dendritic cells
Open accessAbstract
Adoptive transfer of dendritic cells (DCs) transfected with in vitro–transcribed, RNA-encoding, tumor-associated antigens has recently entered clinical testing as a promising approach for cancer immunotherapy. However, pharmacokinetic exploration of RNA as a potential drug compound and a key aspect of clinical development is still pending. While investigating the impact of different structural modifications of RNA molecules on the kinetics of the encoded protein in DCs, we identified components located 3′ of the coding region that contributed to a higher transcript stability and translational efficiency. With the use of quantitative reverse transcription–polymerase chain reaction (RT-PCR) and eGFP variants to measure transcript amounts and protein yield, we showed that a poly(A) tail measuring 120 nucleotides compared with a shorter one, an unmasked poly(A) tail with a free 3′ end rather than one extended with unrelated nucleotides, and 2 sequential β-globin 3′ untranslated regions cloned head to tail between the coding region and the poly(A) tail each independently enhanced RNA stability and translational efficiency. Consecutively, the density of antigen-specific peptide/MHC complexes on the transfected cells and their potency to stimulate and expand antigen-specific CD4+ and CD8+ T cells were also increased. In summary, our data provide a strategy for optimizing RNA-transfected DC vaccines and a basis for defining release criteria for such vaccine preparations.
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Claims the paper makes
17 of 51 cited sources checked · 6 supported · 0 flagged · accuracy not yet measured
- Premisecitation
Similar time kinetics have been reported for intradermally injected DCs in melanoma patients.
“Similar time kinetics have been reported for intradermally injected DCs in melanoma patients.52,53”
Quote in the paperNot a substantial claim2 of 2 sources checked- cites [52] de Vries IJ, Krooshoop DJ, Scharenborg NM, et al. Effective migration of antigen-pulsed dendritic cells to lymph nodes in melanoma patients is determined by their maturation state. Cancer Res. 2003;63: 12-17.
- cites [53] Adema GJ, de Vries IJ, Punt CJ, Figdor CG. Migration of dendritic cell based cancer vaccines: in vivo veritas? Curr Opin Immunol. 2005;17: 170-174. source
The ‘Migration of ex vivo generated DC vaccines’ section reports that intradermally administered DCs reach draining lymph nodes, and the reference annotation for de Vries et al. identifies melanoma patients. Neither passage gives the time course of that migration, so the claimed similarity in time kinetics is unsupported by this copy. (LLM, conf=0.84)
Experimental — accuracy not yet measured
- Premisecitation
d2eGFP has a half-life of 2 hours because of a C-terminal PEST region that promotes protein degradation.
“This variant has a half-life of 2 hours because of a C-terminal PEST region that promotes protein degradation.”
Quote in the paperSubstantial claim1 of 1 sources checked- cites [22] Li X, Zhao X, Fang Y, et al. Generation of destabilized green fluorescent protein as a transcription reporter. J Biol Chem. 1998;273: 34970-34975. source
- Findingown data
Factors involved in RNA stability are induced in the course of maturation of human DCs.
“Notably, by revisiting our database of genomewide expression profiles of immature and mature human DCs obtained with microarrays, 46 we found that factors involved in RNA stability are induced in the course of maturation (data not shown).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [46] Tureci O, Bian H, Nestle FO, et al. Cascades of transcriptional induction during dendritic cell maturation revealed by genome-wide expression analysis. FASEB J. 2003;17: 836-847. source
- Findingown data
For the two beta-globin 3' UTR modification, translational characteristics influenced in immature DCs (primarily message stability and functional half-life) differ from those in mature DCs (primarily translation efficiency).
“In addition, for this structural modification, the translational characteristics influenced in immature DCs (primarily message stability and functional half-life) differed from those in mature DCs (primarily translation efficiency).”
Quote in the paperSubstantial claim - Findingreasoning
Antigen titration shows a higher impact of stabilized IVT RNA constructs on CD4+ compared with CD8+ immune responses.
“Antigen titration shows a higher impact of stabilized IVT RNA constructs on CD4 ϩ compared with CD8 ϩ immune responses, most likely because of the preexistence of highaffinity CD8 ϩ T cells against immunodominant epitopes in the respective donor repertoires.”
Quote in the paperNot a substantial claim
- Findingown data
Density and persistence of peptide/MHC complexes on the cell surface correlate with the amount of antigen-encoding RNA electroporated into cells.
“Density and persistence of peptide/MHC complexes correlated with the amount of RNA transferred into the cells.”
Quote in the paperSubstantial claim - Findingown data
The level and stability of d2eGFP RNA and protein amounts are higher in immature DCs than in mature DCs.
“Interestingly, the level and stability of d2eGFP RNA and the protein amounts were higher in immature than in mature DCs.”
Quote in the paperNot a substantial claim - Findingown data
Inclusion of the human beta-globin 5' UTR does not have an additional beneficial effect on protein yield.
“Notably, inclusion of the human -globin 5Ј UTR did not have an additional beneficial effect on protein yield (data not shown).”
Quote in the paperNot a substantial claim - Findingown data
Encoding the poly(A) tract in the template vector rather than attaching it by enzymatic polyadenylation achieves pure preparations with defined and uniform tail lengths of IVT RNA.
“As we show, this can be achieved by encoding the poly(A) tract in the template vector rather than attaching it by enzymatic polyadenylation.”
Quote in the paperSubstantial claim - Findingown data
The combination of these structural features results in higher protein levels sustained over a longer time, including for molecules with short half-lives.
“In flow cytometric time kinetics with the short-lived d2eGFP variant, we observed that the combination of the respective structural features resulted not only in higher protein levels but was sustained over a longer time and that this applied to molecules with short half-lives (Figure 6C).”
Quote in the paperSubstantial claim - Findingown data
Poly(A)120-tailed RNA yields higher protein levels than shorter poly(A) tailed RNA.
“Importantly, however, higher protein levels resulted in poly(A) 120 -tailed RNA than in shorter RNA.”
Quote in the paperSubstantial claim - Findingown data
In vivo expansion of antigen-specific TCR transgenic CD8+ T cells is significantly superior using Sec-SIINFEKL-2β-globinUTR-A(120) RNA for antigen delivery compared with Sec-SIINFEKL-A(67)ACUAG RNA.
“We found in vivo expansion of antigen-specific TCR transgenic CD8 ϩ T cells to be significantly superior using Sec-SIINFEKL-2-globinUTR-A(120) RNA for antigen delivery compared with Sec-SIINFEKL-A(67)ACUAG RNA (Figure 7B).”
Quote in the paperSubstantial claim - Findingown data
According to our data, the pharmacokinetic properties of RNA in DCs can be further improved by a poly(A) tail 120 bp long.
“According to our data, the pharmacokinetic properties of RNA in DCs can be further improved by a poly(A) tail 120 bp long.”
Quote in the paperSubstantial claim - Findingreasoning
Enzymatically polyadenylated RNA samples generated in independent production runs are not directly comparable.
“Accordingly, enzymatically polyadenylated RNA samples generated in independent production runs were not directly comparable.”
Quote in the paperSubstantial claim - Findingreasoning
With a SIINFEKL-encoding RNA combining all structural improvements, more than 56 pmol SIINFEKL/MHC complexes are expressed on the antigen-presenting cell up to 36 hours after electroporation.
“In line with this, with a SIINFEKLencoding RNA combining all structural improvements we described, more than 56 pmol SIINFEKL/MHC complexes (having been reported as the amount required for induction of robust IFN-␥ secretion of specific T cells in this particular system 17 ) are expressed on the antigen-presenting cell up to 36 hours after electroporation.”
Quote in the paperSubstantial claim - Findingown data
Combining poly(A)120, a free-ending poly(A) tail, and two tandem beta-globin 3' UTRs in one RNA molecule significantly improves RNA stability by approximately 2 orders of magnitude compared to standard IVT RNA with masked poly(A)67 and no UTR.
“Quantification of the transcript in immature DCs by real-time RT-PCR 48 hours after RNA transfer disclosed that combining these modifications significantly improved RNA stability by approximately 2 orders of magnitude (Figure 6A).”
Quote in the paperSubstantial claim - Findingown data
Poly(A) tails longer than 120 bases do not have a significant effect on eGFP or d2eGFP expression.
“Poly(A) tails longer than 120 bases did not have a significant effect on eGFP or d2eGFP expression (data not shown).”
Quote in the paperNot a substantial claim - Findingown data
Prolonged stability from the combination is particularly prominent in immature DCs, with significant MFIs even approximately 80 hours after electroporation.
“Again, prolonged stability was particularly prominent in immature DCs (Figure 6C) in which we observed significant MFIs, even approximately 80 hours after electroporation.”
Quote in the paperSubstantial claim - Findingown data
According to our data, 2 serial 3' UTRs of the β-globin gene upstream of the poly(A) tail are better than one at improving the translational characteristics of RNA.
“According to our data, 2 serial 3Ј UTRs of the -globin gene upstream of the poly(A) tail are better than one at improving the translational characteristics of RNA.”
Quote in the paperSubstantial claim - Findingreasoning
Stability and decay kinetics of the RNAs differ in immature and mature human DCs.
“Interestingly, we observed that stability and decay kinetics of the RNAs differ in immature and mature human DCs, most likely because of differences in the complex network of RNA/protein interactions in both cell types.”
Quote in the paperNot a substantial claim - Findingown data
Protein yield is higher for eGFP IVT RNA variants than for d2eGFP RNA species.
“As expected, protein yield was higher for eGFP IVT RNA variants than for d2eGFP RNA species (Figure 5C).”
Quote in the paperSubstantial claim - Findingown data
RNA with one beta-globin 3' UTR does not substantially increase protein yield compared to tailed control RNA lacking a 3' UTR.
“Interestingly, increased protein yields, particularly in immature DCs, were not as pronounced as expected when comparing RNA with one -globin 3Ј UTR to the tailed control RNA lacking a 3Ј UTR (Figure 5D).”
Quote in the paperSubstantial claim - Study design and conductcitation
Protein decay kinetics is a state-of-the-art tool favored over RNA kinetics for assessing RNA stability because it allows integrated recording of all relevant translational characteristics.
“Protein decay kinetics is regarded as a state-of-the-art tool favored over RNA kinetics because it allows integrated recording of all relevant translational characteristics in the function-oriented manner required for the objective of our investigations.20”
Quote in the paperSubstantial claim0 of 2 sources checked- cites [19] Kren BT, Trembley JH, Steer CJ. Alterations in mRNA stability during rat liver regeneration. Am J Physiol. 1996;270: G763-G777. source
- cites [20] Tanguay RL, Gallie DR. Translational efficiency is regulated by the length of the 3' untranslated region. Mol Cell Biol. 1996;16: 146-156. source
Checked before — its earlier verdict is held back until a new check runs.
- Interpretationown data
Our findings also apply for the expansion of human antigen-specific T cells, as exemplified by stimulation of sorted T-cell populations from CMV-seropositive donors with pp65 antigen.
“Our findings also apply for the expansion of human antigenspecific T cells, as exemplified by stimulation of sorted T-cell populations from CMV-seropositive donors with pp65 antigen.”
Quote in the paperSubstantial claim - Interpretationown data
The optimal poly(A) length, a free-ending poly(A) tail, and an optimized 3' UTR act synergistically on RNA stability and translational efficiency.
“The combination of the optimal poly(A) length, a free-ending poly(A) tail, and an optimized 3 UTR act synergistically on RNA stability and translational efficiency”
Quote in the paperSubstantial claim - Interpretationreasoning
Using type IIS endonucleases for linearization of vectors downstream of the poly(A) stretch prevents an overhang extending the poly(A) tail, which may compromise efficient protein synthesis of the encoded antigen.
“This offers the advantage of cutting within the poly(A) stretch preventing an overhang extending the poly(A) tail, which may compromise efficient protein synthesis of the encoded antigen.”
Quote in the paperSubstantial claim - Premisecitation
Use of RNA results in efficient induction of antigen-specific immune responses in vitro and in animal models.
“Reports demonstrated convincingly that the use of RNA results in efficient induction of antigen-specific immune responses in vitro and in animal models 1,[3][4][5][6][7] and paved the way for trials in humans.”
Quote in the paperSubstantial claim0 of 1 sources checked- cites 1,[3][4][5][6][7]
Couldn’t be matched to a paper in Crossref, so it can’t be checked.
- Interpretationreasoning
The contribution of DriPs to MHC-associated epitopes increases with the rate of de novo translation, which is a function of stability and translational efficiency of the RNA molecule.
“The latter will obviously increase with the rate of de novo translation, which again is a function of stability and translational efficiency of the RNA molecule.”
Quote in the paperSubstantial claim - Interpretationreasoning
Prolonging the duration of antigen presentation may have a significant effect on the stimulation of immune effectors in the lymph node.
“Thus, prolonging the duration of antigen presentation may have a significant effect on the stimulation of immune effectors in the lymph node.”
Quote in the paperSubstantial claim- Rests on an assumption: The capacity of an antigen-encoding IVT RNA vaccine to induce efficient T-cell responses depends on prolonged surface presentation of sufficient amounts of MHC-bound peptide epitopes derived from the RNA-encoded antigen.
- Interpretationreasoning
The difference in protein yield between free-ending and extended poly(A) tail RNA can be accounted for by higher translational efficiency of RNA with a free poly(A) tail.
“Apparently, the difference in protein yield can be accounted for by the higher translational efficiency (eg, the slope of the curve) of RNA with a free poly(A) tail compared with one with a 3Ј extension.”
Quote in the paperSubstantial claim - Interpretationcitation
The effect of 2 serial 3' UTRs on translational characteristics is not mediated simply by the increase of the 3' noncoding region.
“Therefore, we do not expect this effect to be mediated simply by the increase of the 3Ј noncoding region.”
Quote in the paperSubstantial claim
- Classified as probably not a substantial claim. 20
- May combine two claims. 24
- The quote only partly states this claim. 2
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationAntigen-encoding RNA constructs used currently for preclinical studies have poly(A) tails measuring 30 to 70 nucleotides, whereas those administered in clinical trials measure 64 nucleotides.
“Antigen-encoding RNA constructs used currently for preclinical studies have poly(A) tails measuring 30 to 70 nucleotides in length, 4,5 whereas those administered in clinical trials measure 64 nucleotides in length.8,12”
Quote in the paperNot a substantial claim2 of 4 sources checked- cites [4] van Tendeloo VF, Ponsaerts P, Lardon F, et al. Highly efficient gene delivery by mRNA electroporation in human hematopoietic cells: superiority to lipofection and passive pulsing of mRNA and to electroporation of plasmid cDNA for tumor antigen loading of dendritic cells. Blood. 2001;98: 49-56. source
The ‘Production of in vitro-transcribed mRNA’ section describes the RNA templates and transcription method but gives no poly(A)-tail length. Searches for 64-nucleotide and poly(A) tails found no passage addressing the length of RNA constructs administered in clinical trials. (LLM, conf=0.76)
Experimental — accuracy not yet measured - cites [5] Teufel R, Carralot JP, Scheel B, et al. Human peripheral blood mononuclear cells transfected with messenger RNA stimulate antigen-specific cytotoxic T-lymphocytes in vitro. Cell Mol Life Sci. 2005;62: 1755-1762. source
- cites [8] Su Z, Dannull J, Yang BK, et al. Telomerase mRNA-transfected dendritic cells stimulate antigen-specific CD8+ and CD4+ T cell responses in patients with metastatic prostate cancer. J Immunol. 2005;174: 3798-3807. source
- cites [12] Heiser A, Coleman D, Dannull J, et al. Autologous dendritic cells transfected with prostate-specific antigen RNA stimulate CTL responses against metastatic prostate tumors. J Clin Invest. 2002; 109: 409-417. source
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationThe poly(A) tail regulates the fate of mRNA by synergy with the 5' cap, the internal ribosomal entry site, and various RNA stability determinants.
“The poly(A) tail regulates the fate of mRNA by synergy with the 5Ј cap, with the internal ribosomal entry site, and with various RNA stability determinants.31,[38][39][40][41]”
Quote in the paperNot a substantial claim2 of 5 sources checked- cites [31] Gallie DR. A tale of two termini: a functional interaction between the termini of an mRNA is a pre-requisite for efficient translation initiation. Gene. 1998;216: 1-11. source
- cites [38] Grosset C, Chen CY, Xu N, et al. A mechanism for translationally coupled mRNA turnover: interaction between the poly(A) tail and a c-fos RNA coding determinant via a protein complex. Cell. 2000;103: 29-40. source
The passages labeled Abstract (page unavailable) describe an interaction between the poly(A) tail and 5′ cap that influences translation, and experiments linking the tail to an mRNA decay determinant, the mCRD. They also mention cap-independent initiation, but do not establish that the poly(A) tail acts synergistically with an internal ribosomal entry site. (LLM, conf=0.84)
Experimental — accuracy not yet measured - cites [39] Wakiyama M, Imataka H, Sonenberg N. Interaction of eIF4G with poly(A)-binding protein stimulates translation and is critical for Xenopus oocyte maturation. Curr Biol. 2000;10: 1147-1150. source
- cites [40] Wang Z, Day N, Trifillis P, Kiledjian M. An mRNA stability complex functions with poly(A)-binding protein to stabilize mRNA in vitro. Mol Cell Biol. 1999;19: 4552-4560. source
The Discussion (page not available in our copy) describes an interaction between a stability complex on the α-globin mRNA 3′ UTR and poly(A)-binding protein that may slow deadenylation. The paper also discusses protective roles for the 5′ cap and poly(A) tail, but the passages do not establish synergy between them or address synergy with an internal ribosomal entry site. (LLM, conf=0.86)
Experimental — accuracy not yet measured - cites [41] Preiss T, Muckenthaler M, Hentze MW. Poly(A)-tail-promoted translation in yeast: implications for translational control. RNA. 1998;4: 1321-1331. source
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationRegulation of mRNA turnover plays a major role in the overall control of gene expression, and the mechanisms underlying steady-state levels of naturally expressed mRNAs in the cytoplasm have received considerable attention.
“The mechanisms underlying the steady state levels of naturally expressed mRNAs in the cytoplasm have received considerable attention recently because the regulation of mRNA turnover plays a major role in the overall control of gene expression.26,27”
Quote in the paperNot a substantial claim1 of 2 sources checked- cites [26] Gebauer F, Hentze MW. Molecular mechanisms of translational control. Nat Rev Mol Cell Biol. 2004;5: 827-835. source
The Main section discusses translational control of gene expression, including whether cytoplasmic mRNAs are actively translated. It does not address mRNA turnover as a determinant of steady-state mRNA levels or say that those mechanisms have received considerable attention (searched: mRNA turnover, steady-state levels, stability, degradation and decay). (LLM, conf=0.72)
Experimental — accuracy not yet measured - cites [27] Cleveland DW. Gene regulation through messenger RNA stability. Curr Opin Cell Biol. 1989;1: 1148-1153. source
May combine two claims.
PremisecitationAntigen-encoding RNA has the advantages of a genetic vaccine (delivery of all epitopes of the whole antigen, easy manufacturing, standardized purification) and is a safe pharmaceutical characterized by transient expression and lack of integration into the genome of the treated host.
“Antigen-encoding RNA 1,2 has the advantages of a genetic vaccine (delivery of all epitopes of the whole antigen, easy manufacturing, standardized purification) and the added value of a safe pharmaceutical characterized by transient expression and lack of integration into the genome of the treated host.”
Quote in the paperSubstantial claim1 of 2 sources checked- cites [1] Hoerr I, Obst R, Rammensee HG, Jung G. In vivo application of RNA leads to induction of specific cytotoxic T lymphocytes and antibodies. Eur J Immunol. 2000;30: 1-7. source
- cites [2] Carralot JP, Probst J, Hoerr I, et al. Polarization of immunity induced by direct injection of naked sequence-stabilized mRNA vaccines. Cell Mol Life Sci. 2004;61: 2418-2424. source
Split into 2 atoms — 1 confirmed, 1 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.
Experimental — accuracy not yet measured
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationThe length of the 3' UTR has an effect on the stability and translational efficiency of poly(A)-negative RNA but not of poly(A)-positive RNA, for which a 27-bp 3' UTR appears to be optimal.
“As previous reports show, the length of the 3Ј UTR as such has an effect on the stability and translational efficiency of poly(A)-negative RNA but not of poly(A)-positive RNA, for which a 27-bp 3Ј UTR appears to be optimal.20”
Quote in the paperNot a substantial claim1 of 1 sources checked- cites [20] Tanguay RL, Gallie DR. Translational efficiency is regulated by the length of the 3' untranslated region. Mol Cell Biol. 1996;16: 146-156. source
The Results report that increasing 3′ UTR length increased translational efficiency and half-life for poly(A)-negative mRNA, while these measures varied little for poly(A)-positive mRNA over most lengths tested. The claim's categorical 'not' misses a substantial exception: a 7-base UTR sharply reduced translation of poly(A)-positive mRNA. The reported 27-base optimum is specifically for poly(A)-tail-mediated stimulation of translation. Page numbers are not provided in the supplied passages. (LLM, conf=0.88)
Experimental — accuracy not yet measured
Classified as probably not a substantial claim.
PremisecitationThe amount of SIINFEKL/MHC complexes required for induction of robust IFN-γ secretion of specific T cells in this particular system is 56 pmol.
“(having been reported as the amount required for induction of robust IFN-␥ secretion of specific T cells in this particular system 17 )”
Quote in the paperSubstantial claim1 of 1 sources checked- cites [17] Porgador A, Yewdell JW, Deng Y, Bennink JR, Germain RN. Localization, quantitation, and in situ detection of specific peptide-MHC class I complexes using a monoclonal antibody. Immunity. 1997;6: 715-726. source
In “Sensitivity of Ligand Detection” (page unavailable), the paper compares SIINFEKL concentrations with IFN-γ secretion by a specific CD8+ T-cell line. Our copy discusses minimal concentrations for measurable secretion, but does not establish 56 pmol as an amount required for robust secretion; it reports peptide concentrations in pM, not amounts in pmol. (LLM, conf=0.86)
Experimental — accuracy not yet measured
Probably not a substantial claim: may not pass S3 (load-bearing).
InterpretationreasoningThe improvement of RNA kinetics by these modifications may also be of interest for other cellular settings, such as T cells, B cells, and muscle cells.
“In principle, the improvement of RNA kinetics by these modifications is not restricted to DCs but is similar in K562 cells and may also be of interest for other cellular settings, such as T cells, 43 B cells, 44 and muscle cells, 45 which are attractive host cells for the expression of IVT RNA-encoded proteins.”
Quote in the paperNot a substantial claim2 of 3 sources checked- cites [43] Zhao Y, Zheng Z, Cohen CJ, et al. High-efficiency transfection of primary human and mouse T lymphocytes using RNA electroporation. Mol Ther. 2006;13: 151-159. source
- cites [44] Van den Bosch GA, Ponsaerts P, Nijs G, et al. Ex vivo induction of viral antigen-specific CD8 T cell responses using mRNA-electroporated CD40-activated B cells. Clin Exp Immunol. 2005;139: 458-467. source
- cites [45] Wolff JA, Malone RW, Williams P, et al. Direct gene transfer into mouse muscle in vivo. Science. 1990;247: 1465-1468. source
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationVectors used currently for preclinical and clinical studies resort either to UTRs of β-globin or to autochthonous UTRs of the antigen they deliver, which are not further characterized in terms of their impact on RNA stability.
“Vectors used currently for preclinical and clinical studies resort either to UTRs of -globin 5,14,23 or to autochthonous UTRs of the antigen they want to deliver, 12 which are not further characterized in terms of their impact on RNA stability.”
Quote in the paperNot a substantial claim1 of 4 sources checked- cites [5] Teufel R, Carralot JP, Scheel B, et al. Human peripheral blood mononuclear cells transfected with messenger RNA stimulate antigen-specific cytotoxic T-lymphocytes in vitro. Cell Mol Life Sci. 2005;62: 1755-1762. source
The Messenger RNA section (page unavailable) says the constructs used in this study contain UTRs from globin mRNAs, supporting this paper as an example of globin-UTR use. It does not specify β-globin or characterize the UTRs’ impact on RNA stability. (LLM, conf=0.82)
Experimental — accuracy not yet measured - cites [14] Michiels A, Tuyaerts S, Bonehill A, et al. Electroporation of immature and mature dendritic cells: implications for dendritic cell-based vaccines. Gene Ther. 2005;12: 772-782. source
- cites [23] Conry RM, LoBuglio AF, Wright M, et al. Characterization of a messenger RNA polynucleotide vaccine vector. Cancer Res. 1995;55: 1397-1400.
- cites [12] Heiser A, Coleman D, Dannull J, et al. Autologous dendritic cells transfected with prostate-specific antigen RNA stimulate CTL responses against metastatic prostate tumors. J Clin Invest. 2002; 109: 409-417. source
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationSequence elements within the 3' and 5' noncoding regions of RNA have been characterized as important factors in regulating transcript stability, in ways that are not completely understood.
“Sequence elements within the 3Ј and 5Ј noncoding regions of the RNA molecule have been characterized as important factors in regulating the stability of a transcript in ways that are not completely understood.32,33”
Quote in the paperNot a substantial claim2 of 2 sources checked- cites [32] Mignone F, Gissi C, Liuni S, Pesole G. Untranslated regions of mRNAs. Genome Biol. 2002;3: Reviews0004. source
- cites [33] Yu J, Russell JE. Structural and functional analysis of an mRNP complex that mediates the high stability of human beta-globin mRNA. Mol Cell Biol. 2001;21: 5879-5888. source
The paper’s introductory discussion says that cis-acting elements, usually in the 3′ UTR, regulate mRNA stability; the Discussion identifies a stability element in the β-globin 3′ UTR. It also describes the 5′ cap as a general stabilizing feature, but does not characterize sequence elements within the 5′ noncoding region as stability regulators. Page numbers are not provided in our copy. (LLM, conf=0.86)
Experimental — accuracy not yet measured
May combine two claims.
PremisecitationMouse DCs start to enter the regional lymph node 18 hours after inoculation in the skin, reaching a maximum within 72 hours.
“Mouse DCs start to enter the regional lymph node 18 hours after inoculation in the skin, reaching a maximum within 72 hours.50,51”
Quote in the paperSubstantial claim1 of 2 sources checked- cites [50] Martin-Fontecha A, Sebastiani S, Hopken UE, et al. Regulation of dendritic cell migration to the draining lymph node: impact on T lymphocyte traffic and priming. J Exp Med. 2003;198: 615-621. source
In Results and Discussion, the authors report recovering injected mouse DCs from draining lymph nodes on day 2. They do not establish that entry begins at 18 hours or that DC numbers peak within 72 hours. The plateau by days 3–4 refers to total lymph-node cellularity, not DC migration. (LLM, conf=0.84)
Experimental — accuracy not yet measured - cites [51] Randolph GJ, Angeli V, Swartz MA. Dendritic-cell trafficking to lymph nodes through lymphatic vessels. Nat Rev Immunol. 2005;5: 617-628. source
May combine two claims. The quote only partly states this claim.
Findingown dataExpansion of T cells measured on day 7 in an IFN-γ ELISpot using autologous DCs pulsed with pp65 peptides showed superiority of Sec-pp65-2β-globinUTR-A(120) over Sec-pp65-A(67)ACUAG, with effects most prominent for CD4+ T-cell expansion.
“Expansion of T cells measured on day 7 in an IFN-␥ ELISpot using autologous DCs pulsed with a pool of overlapping peptides covering either the entire pp65 protein sequence or a control protein showed superiority of Sec-pp65-2-globinUTR-A(120), with effects most prominent for the expansion of CD4 ϩ T cells.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataAfter electroporation of as little as 10 pmol RNA, surface peptide/MHC expression is detectable at 12 hours, declines over time, and is below detection at 24 hours.
“Even after electroporation of RNA amounts as low as 10 pmol, surface expression of peptide/MHC molecules was easily detected 12 hours after transfection, declined over time, and was below the detection level 24 hours after RNA transfer (Figure 2).”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataMessage stability and functional RNA half-life are comparable for eGFP-A(67) and eGFP-A(67)ACUAG RNAs.
“Interestingly, visual inspection of the curves showed that the message stability (eg, the length of time over which protein continued to accumulate) of both eGFP-A(67) and eGFP-A(67)ACUAG RNAs appeared to be comparable (Figure 4B).”
- Also stated in: § Overhang at the 3 end of the poly(A) tail hampers translational efficiency of IVT RNA · plain
“In addition, functional RNA half-life (eg, time needed to complete a 50% decay in the capacity of an RNA to synthesize protein, 21 which is a more accurate parameter than physical half-life because it also considers translational competence of an RNA species) is comparable for eGFP-A(67) and eGFP-A(67)ACUAG RNAs.”
Quote in the paperNot a substantial claim- Also stated in: § Overhang at the 3 end of the poly(A) tail hampers translational efficiency of IVT RNA · plain
May combine two claims.
Findingown dataProtein levels are higher and decay is protracted in immature DCs compared with mature DCs, with maximum protein levels sustained in steady state until 120 hours after electroporation.
“Again, protein levels were higher and decay was protracted in immature DCs compared with mature ones, with maximum protein levels sustained in steady state until 120 hours after electroporation (Figure 5C).”
Quote in the paperNot a substantial claimMay combine two claims.
Findingown dataRNA with two sequential beta-globin 3' UTRs between the coding sequence and the poly(A) tail results in significantly higher maximum protein levels and prolonged persistence compared to control.
“However, 2 reiterated -globin 3Ј UTRs between the coding sequence and the poly(A) tail resulted in significantly higher maximum protein levels and prolonged persistence of the protein.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataCombining the structural components achieved a more than 5-fold increase of maximum protein concentration and persistence of protein expression over a longer time.
“By combining these structural components, we achieved not only a more than 5-fold increase of maximum protein concentration but also persistence of protein expression over a longer time.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataThe RNA modifications increase the density of antigen-specific peptide/MHC complexes on transfected cells and their potency to stimulate and expand antigen-specific CD4+ and CD8+ T cells.
“Consecutively, the density of antigen-specific peptide/MHC complexes on the transfected cells and their potency to stimulate and expand antigen-specific CD4 ؉ and CD8 ؉ T cells were also increased.”
- Also stated in: § Introduction · boosted
“We show that this optimization improves the surface presentation of epitopes and the induction of T-cell responses.”
Quote in the paperSubstantial claim- Also stated in: § Introduction · boosted
May combine two claims.
Findingown dataRNA with a free-ending poly(A) tail yields 1.5-fold higher maximum protein levels and prolonged detectability compared with RNA with a 3' extension in human immature DCs.
“Linearization with Sap1 improved expression significantly compared with restriction with Spe1 because the RNA with free-ending poly(A) tail resulted in 1.5-fold higher maximum protein levels and prolonged detectability of the protein.”
Quote in the paperSubstantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
Findingnone givenThe improvement of RNA kinetics by these modifications is similar in K562 cells.
“In principle, the improvement of RNA kinetics by these modifications is not restricted to DCs but is similar in K562 cells and may also be of interest for other cellular settings, such as T cells, 43 B cells, 44 and muscle cells, 45 which are attractive host cells for the expression of IVT RNA-encoded proteins.”
Quote in the paperNot a substantial claimMay combine two claims.
Findingown dataPoly(A)120-tailed RNA has higher intracellular RNA levels than shorter poly(A) tailed RNA, suggesting shorter tails undergo faster degradation.
“As exemplified for immature and mature DCs by real-time RT PCR analysis, intracellular IVT RNA levels of d2eGFP variants at different time points after transfection were highest for poly(A) 120tailed RNA, suggesting that RNAs with shorter poly(A) tails undergo faster degradation (Figure 5A).”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataThe optimized RNA (Sec-SIINFEKL-2beta-globinUTR-A(120)) yields significantly higher numbers of OVA peptide/Kb complexes on the cell surface sustained over a longer time course compared to standard RNA upon electroporation into EL4 cells and BMDCs.
“We detected significantly higher numbers of OVA peptide/K b complexes on the cell surface sustained over a longer time course upon electroporation of the genetically improved RNA Sec-SIINFEKL-2-globinUTR-A(120) (Figure 7A).”
Quote in the paperSubstantial claimMay combine two claims. The quote only partly states this claim.
Findingown dataEnzymatic polyadenylation of RNA yields a mixture of transcripts varying in poly(A) tail length, and the number of attached ATP nucleotides cannot be precisely controlled even under the same reaction conditions.
“Because many parameters influenced poly(A) tail length during the polyadenylation reaction, the number of attached ATP nucleotides could not be precisely controlled even if the same reaction conditions were applied.”
- Also stated in: § Enzymatic polyadenylation of RNA results in a mixture of transcripts varying in length · hedged
“In addition to this variability between different preparations, each individual preparation appeared to contain a mixture of RNA species differing in poly(A) tail length.”
Quote in the paperSubstantial claim- Also stated in: § Enzymatic polyadenylation of RNA results in a mixture of transcripts varying in length · hedged
May combine two claims.
InterpretationcitationThe observations that MHC epitope density correlates with intracellular protein and that epitope production occurs without detectable full-length protein are not contradictory and may be explained by defective ribosomal products (DriPs) being quickly processed into peptides and contributing significantly to MHC-associated epitopes.
“These observations are not contradictory but may be explained by the fact that in addition to steady state protein levels, defective ribosomal products (DriPs) such as unfinished or misfolded proteins are quickly processed into peptides and contribute significantly to MHC-associated epitopes.49”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [49] Yewdell JW, Anton LC, Bennink JR. Defective ribosomal products (DRiPs): a major source of antigenic peptides for MHC class I molecules? J Immunol. 1996;157: 1823-1826. source
Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).
Interpretationnone givenOne of the advantages of formulating the antigen as IVT RNA is the added value provided by recombinant DNA technology of easily engineering elements into vaccine constructs that enhance their potency.
“Moreover, our report underscores that one of the many advantages of formulating the antigen as IVT RNA is the added value provided by recombinant DNA technology of easily engineering elements into vaccine constructs that enhance their potency.”
Quote in the paperNot a substantial claimClassified as probably not a substantial claim.
PremisecitationProduction of MHC-associated epitopes and priming of cytotoxic T-lymphocytes is achieved even if the full-length protein encoded by the transferred RNA cannot be detected in DCs.
“However, it has also been reported that the production of MHC-associated epitopes and priming of cytotoxic T-lymphocytes is achieved even if the full-length protein encoded by the transferred RNA cannot be detected in DCs.42,48”
Quote in the paperSubstantial claim0 of 2 sources checked- cites [42] Boczkowski D, Nair SK, Snyder D, Gilboa E. Dendritic cells pulsed with RNA are potent antigen-presenting cells in vitro and in vivo. J Exp Med. 1996;184: 465-472. source
Checked before — its earlier verdict is held back until a new check runs.
- cites [48] Grunebach F, Muller MR, Nencioni A, Brossart P. Delivery of tumor-derived RNA for the induction of cytotoxic T-lymphocytes. Gene Ther. 2003;10: 367-374. source
Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).
InterpretationreasoningThe higher impact of stabilized IVT RNA constructs on CD4+ compared with CD8+ immune responses is most likely because of the preexistence of high-affinity CD8+ T cells against immunodominant epitopes in the respective donor repertoires.
“Antigen titration shows a higher impact of stabilized IVT RNA constructs on CD4 ϩ compared with CD8 ϩ immune responses, most likely because of the preexistence of highaffinity CD8 ϩ T cells against immunodominant epitopes in the respective donor repertoires.”
Quote in the paperNot a substantial claimMay combine two claims.
Interpretationown dataTemplate-encoded poly(A) tails reproducibly provide a well-defined product of sufficient purity and are superior to enzymatic polyadenylation.
“In summary, templateencoded poly(A) tails were clearly superior because they reproducibly provided a well-defined product of sufficient purity.”
Quote in the paperSubstantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationThe density of MHC epitopes presented on the cell surface correlates with the amount of intracellular protein detected at a defined time point.
“Previous studies reported that the density of MHC epitopes presented on the cell surface correlates with the amount of intracellular protein detected at a defined time point.47”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [47] Princiotta MF, Finzi D, Qian SB, et al. Quantitating protein synthesis, degradation, and endogenous antigen processing. Immunity. 2003;18: 343-354. source
Checked before — its earlier verdict is held back until a new check runs.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationIn some reports, the poly(A) tail is attached enzymatically by terminal polyadenylation instead of being encoded in the template.
“In some reports, the poly(A) tail is attached enzymatically by terminal polyadenylation instead of being encoded in the template.13”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [13] Steitz J, Britten CM, Wolfel T, Tuting T. Effective induction of anti-melanoma immunity following genetic vaccination with synthetic mRNA coding for the fusion protein EGFP.TRP2. Cancer Immunol Immunother. 2006;55: 246-253. source
May combine two claims.
PremisecitationEarly clinical trials of RNA-based vaccines showed feasibility, lack of toxicity, and promising efficacy based on immunologic and clinical read-outs.
“Early clinical trials showed feasibility, lack of toxicity, and promising efficacy based on immunologic and clinical read-outs.[8][9][10][11][12]”
Quote in the paperNot a substantial claim0 of 5 sources checked- cites [8] Su Z, Dannull J, Yang BK, et al. Telomerase mRNA-transfected dendritic cells stimulate antigen-specific CD8+ and CD4+ T cell responses in patients with metastatic prostate cancer. J Immunol. 2005;174: 3798-3807. source
- cites [9] Caruso DA, Orme LM, Neale AM, et al. Results of a phase 1 study utilizing monocyte-derived dendritic cells pulsed with tumor RNA in children and young adults with brain cancer. Neuro-oncol. 2004;6: 236-246. source
Checked before — its earlier verdict is held back until a new check runs.
- cites [10] Caruso DA, Orme LM, Amor GM, et al. Results of a phase I study utilizing monocyte-derived dendritic cells pulsed with tumor RNA in children with stage 4 neuroblastoma. Cancer. 2005;103: 1280-1291. source
- cites [11] Morse MA, Nair SK, Mosca PJ, et al. Immunotherapy with autologous, human dendritic cells transfected with carcinoembryonic antigen mRNA. Cancer Invest. 2003;21: 341-349. source
- cites [12] Heiser A, Coleman D, Dannull J, et al. Autologous dendritic cells transfected with prostate-specific antigen RNA stimulate CTL responses against metastatic prostate tumors. J Clin Invest. 2002; 109: 409-417. source
Classified as probably not a substantial claim. May combine two claims.
PremisecitationmRNA stability and translation are controlled by a complex network of RNA/protein interactions and depend on the primary and secondary structures of mRNAs, their rates of translation, and the initiation of one of the various degradation mechanisms.
“mRNA stability and translation are controlled by a complex network of RNA/protein interactions and depend on the primary and secondary structures of mRNAs, their rates of translation, and the initiation of one of the various degradation mechanisms.[28][29][30][31]”
Quote in the paperSubstantial claim0 of 4 sources checked- cites [28] Guhaniyogi J, Brewer G. Regulation of mRNA stability in mammalian cells. Gene. 2001;265: 11-23. source
- cites [29] Preiss T, Hentze W. Starting the protein synthesis machine: eukaryotic translation initiation. Bioessays. 2003;25: 1201-1211. source
- cites [30] Meyer S, Temme C, Wahle E. Messenger RNA turnover in eukaryotes: pathways and enzymes. Crit Rev Biochem Mol Biol. 2004;39: 197-216. source
- cites [31] Gallie DR. A tale of two termini: a functional interaction between the termini of an mRNA is a pre-requisite for efficient translation initiation. Gene. 1998;216: 1-11. source
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
Study design and conductown dataFlow cytometric data in Figure 6C are from 3 independent experiments conducted in parallel, showing nearly no SD and high reproducibility.
“Flow cytometric data presented in Figure 6C feature results of 3 independent experiments conducted in parallel, showing nearly no SD and proving the high reproducibility of this procedure.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
InterpretationreasoningThe difference in stability and decay kinetics of RNAs between immature and mature human DCs is most likely because of differences in the complex network of RNA/protein interactions in both cell types.
“Interestingly, we observed that stability and decay kinetics of the RNAs differ in immature and mature human DCs, most likely because of differences in the complex network of RNA/protein interactions in both cell types.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing), S4 (based).
Interpretationnone givenFor human DCs, this is the only study systematically investigating the influence of several 3' elements on translational characteristics of transiently expressed RNA.
“For human DCs, this is the only study systematically investigating the influence of several 3Ј elements on translational characteristics of transiently expressed RNA.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationIn vitro-transcribed RNA used in such studies typically consists of a 5' cap structure followed by the sequence encoding the antigen of interest and a poly(A) tail 30 to 70 nucleotides in length.
“In vitro-transcribed RNA generated from plasmid templates and used in such studies typically consists of a 5Ј cap structure followed by the sequence encoding the antigen of interest and a poly(A) tail 30 to 70 nucleotides in length.4,5”
Quote in the paperNot a substantial claim0 of 2 sources checked- cites [4] van Tendeloo VF, Ponsaerts P, Lardon F, et al. Highly efficient gene delivery by mRNA electroporation in human hematopoietic cells: superiority to lipofection and passive pulsing of mRNA and to electroporation of plasmid cDNA for tumor antigen loading of dendritic cells. Blood. 2001;98: 49-56. source
Checked before — its earlier verdict is held back until a new check runs.
- cites [5] Teufel R, Carralot JP, Scheel B, et al. Human peripheral blood mononuclear cells transfected with messenger RNA stimulate antigen-specific cytotoxic T-lymphocytes in vitro. Cell Mol Life Sci. 2005;62: 1755-1762. source
Checked before — its earlier verdict is held back until a new check runs.
May combine two claims.
Interpretationown dataThe paper's data provide a strategy for optimizing RNA-transfected DC vaccines and a basis for defining release criteria for such vaccine preparations.
“In summary, our data provide a strategy for optimizing RNA-transfected DC vaccines and a basis for defining release criteria for such vaccine preparations.”
Quote in the paperSubstantial claimProbably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationThe gene of interest is used with its autochthonous noncoding regions or is flanked by the untranslated region of the human or Xenopus laevis beta-globin gene.
“The gene of interest is used with its autochthonous noncoding regions 12 or is flanked by the untranslated region (UTR) of the human or Xenopus laevis -globin gene.14”
Quote in the paperNot a substantial claim0 of 2 sources checked- cites [12] Heiser A, Coleman D, Dannull J, et al. Autologous dendritic cells transfected with prostate-specific antigen RNA stimulate CTL responses against metastatic prostate tumors. J Clin Invest. 2002; 109: 409-417. source
- cites [14] Michiels A, Tuyaerts S, Bonehill A, et al. Electroporation of immature and mature dendritic cells: implications for dendritic cell-based vaccines. Gene Ther. 2005;12: 772-782. source
May combine two claims.
Interpretationown dataOptimization of structural features of antigen-encoding RNA increases the density and stability of peptide/MHC complexes on the cell surface and translates into improved stimulation of CD4+ and CD8+ T cells in murine and human systems.
“In summary, we confirm our starting assumption that the optimization of structural features of antigen-encoding RNA increases the density and stability of peptide/MHC complexes on the cell surface and translates into improved stimulation of CD4 ϩ and CD8 ϩ T cells in murine and human systems.”
Quote in the paperSubstantial claim- Rests on an assumption: The capacity of an antigen-encoding IVT RNA vaccine to induce efficient T-cell responses depends on prolonged surface presentation of sufficient amounts of MHC-bound peptide epitopes derived from the RNA-encoded antigen.
The paper relies on this without support.
Neither the pharmacokinetics of antigen-encoding RNA within DCs nor the impact of structural features of the drug formulation on its pharmacologic properties has thus far been addressed for antigen-encoding RNA.
“Neither of these questions has thus far been addressed for antigen-encoding RNA.”
The paper relies on this without support.
The effects of each structural feature on the intracellular bioavailability of IVT RNA have not been systematically investigated.
“The effects of each of these structural features on the intracellular bioavailability of the RNA molecule have not been systematically investigated.”
The paper relies on this without support.
Little is known about the time-dose curve and the translational efficiency of IVT antigen-encoding RNA after its electroporation into DCs.
“In fact, surprisingly little is known about the time-dose curve and the translational efficiency of in vitro-transcribed (IVT) antigen-encoding RNA after its electroporation into DCs.”
The paper relies on this without support.
A dose-response relationship between the levels of antigen-encoding RNA within the cell and the peptide/MHC molecules on the cell surface has not been established.
“Similarly, a dose-response relationship between the levels of antigen-encoding RNA within the cell and the peptide/MHC molecules on the cell surface has not been established.”
The paper relies on this without support.
The extent to which the density of peptide/MHC complexes on the cell surface can be influenced by augmenting RNA stability remains unclear.
“Thus, the extent to which the density of peptide/ MHC complexes on the cell surface can be influenced by augmenting RNA stability remains unclear.”
The paper relies on this without support.
This is the first study to determine the structural modifications that increase the stability and translational efficiency of RNA molecules in DCs.
“To our knowledge, this is the first study to determine the structural modifications that increase the stability and translational efficiency of RNA molecules in DCs.”
The paper relies on this without support.
The capacity of an antigen-encoding IVT RNA vaccine to induce efficient T-cell responses depends on prolonged surface presentation of sufficient amounts of MHC-bound peptide epitopes derived from the RNA-encoded antigen.
“The mode of action of such a vaccine and its capacity to induce efficient T-cell responses depends on prolonged surface presentation of sufficient amounts of MHC-bound peptide epitopes derived from the RNA-encoded antigen.”
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
Previous strategies to increase mRNA stability have yielded modest improvements in RNA stability.
“These strategies have yielded modest improvements in RNA stability10, 13–16, and thus alternative approaches to RNA stabilization, such as circularization, are desirable.”
From Engineering circular RNA for potent and stable translation in eukaryotic cells (2018) · cites this paper as [13]
In the section on combining poly(A)-tail and 3′ UTR modifications (page unavailable), the paper reports an approximately two-orders-of-magnitude improvement in RNA stability relative to standard IVT RNA. That result is contrary to describing the improvement reported by this source as modest. (LLM, conf=0.88)
Checked by a reasoning model — may be wrong.
Experimental — accuracy not yet measuredA poly(A) tail of 120 units is considered the standard size for mRNA vaccines.
“A poly(A) tail of 120 units is considered the standard size for mRNA vaccines.9,20,21”
From Combining an optimized mRNA template with a double purification process allows strong expression of in vitro transcribed mRNA (2021) · cites this paper as [9]
The Discussion says antigen-encoding RNAs then used in preclinical studies had 30–70-nucleotide poly(A) tails, while those in clinical trials had 64-nucleotide tails. It proposes 120 nucleotides as an improvement, not an established standard; the section on combining modifications calls a 67-base-tail RNA the standard comparator. (LLM, conf=0.88)
Checked by a reasoning model — may be wrong.
Experimental — accuracy not yet measuredBoth BNT162b1 and BNT162b2 are pharmacologically optimized N1-methylpseudouridine nucleoside-modified mRNAs given intramuscularly in a prime-boost regimen.
“Both of the candidates are pharmacologically optimized 15,16 , N 1 -methylpseudouridine nucleoside-modified mRNAs 17 that are administered intramuscularly in a prime-boost regimen 21”
From BNT162b2 vaccine induces neutralizing antibodies and poly-specific T cells in humans (2021) · cites this paper as [15]
The antigen-encoding RNA contains sequence elements that increase RNA stability and translation efficiency in human dendritic cells.
“The antigen-encoding RNA contains sequence elements that increase RNA stability and translation efficiency in human dendritic cells 15,16 .”
From BNT162b2 vaccine induces neutralizing antibodies and poly-specific T cells in humans (2021) · cites this paper as [15]
The steady effect of human β-globin 3′ UTR sequences is enhanced when two human β-globin 3′ UTRs are placed in a head-to-tail orientation.
“The steady effect of human β-globin 3ʹ-UTR sequences is enhanced when two human β-globin 3ʹ-UTRs are placed in a head-to-tail orientation (Holtkamp et al. 2006).”
From Modifications of mRNA vaccine structural elements for improving mRNA stability and translation efficiency (2022) · cites this paper as Holtkamp et al. 2006
The structural characteristics of mRNA such as the CAP, poly(A) tail, and UTRs directly impact mRNA stability and translation efficiency.
“The structural characteristics of mRNA such as the CAP, poly(A) tail, and untranslated regions (UTRs) directly impact mRNA stability and translation efficiency (Orlandini von Niessen et al. 2019; Holtkamp et al. 2006).”
From Modifications of mRNA vaccine structural elements for improving mRNA stability and translation efficiency (2022) · cites this paper as Holtkamp et al. 2006
The design of non-coding structural elements of mRNA such as the CAP structure, poly(A) tail and untranslated regions has a major impact on mRNA stability and translation capacity.
“The design of (non-coding) structural elements of the mRNA such as the CAP structure, poly(A) tail and untranslated regions (UTRs) all have a major impact on the mRNA stability and translation capacity [31,32]”
From The dawn of mRNA vaccines: the COVID-19 case (2021) · cites this paper as [32]
Correlations exist between elongation of the poly(A) tail of mRNA and duration of expression, and specific 3' UTR modifications can slow down decay of the poly(A) tail by deadenylation.
“Correlations were found between the elongation of the poly(A) tail of mRNA and the duration of expression, and specific modifications at the 3' UTR were reported that can slow-down the decay of the poly(A) tail by deadenylation [57,58].”
From Three decades of messenger RNA vaccine development (2019) · cites this paper as [57]
Specific UTRs, such as one or two copies of the 3′ UTR of globin or a short 14-nucleotide 5′ UTR, increase the translational efficiency of mRNAs.
“numerous works have developed specific UTRs to increase the translational efficiency of mRNAs, as the use of one or two copies of the 3′ UTR of the globin or other known stable mRNA8,9 or short 14-nucleotide 5′ UTR.27”
From Combining an optimized mRNA template with a double purification process allows strong expression of in vitro transcribed mRNA (2021) · cites this paper as [9]
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- Authors
- Holtkamp, S., Kreiter, S., Selmi, A., Simon, P., Koslowski, M., Huber, C., Türeci, O., & Sahin, U.
- Affiliations
- Johannes Gutenberg University Mainz (DE)
- From the Department of Internal Medicine III, Division of Experimental and Translational Oncology, Johannes-Gutenberg University, Mainz, Germany.
- Venue
- Blood
- Publisher
- American Society of Hematology
- Published
- 2006-12-15
- Volume
- 108
- Issue
- 13
- Pages
- 4009-4017
- ISSN
- 0006-4971 (ISSN-L)
- 1528-0020
- OpenAlex
- W2039422212
- Semantic Scholar
- 48122d1dd006266c2abd2c302acdc4fe220181de
- Cited by
- 625
- References
- 54
- Citations by year
- 2026: 51 · 2025: 62 · 2024: 60 · 2023: 73 · 2022: 77 · 2021: 62 · 2020: 35 · 2019: 25 · 2018: 35 · 2017: 20 · 2016: 24 · 2015: 22 · 2014: 11 · 2013: 13 · 2012: 7
- FWCI
- 2.23
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Locations
- Blood (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Deutsche Forschungsgemeinschaft
- Topics
- Life Sciences › Immunology and Microbiology › Immunology › Immunotherapy and Immune Responses
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
- Keywords
- dendritic cell vaccine, RNA vaccines, tumor-associated antigens, poly(A) tail length, 3' untranslated region, transcript stability, translation efficiency, antigen presentation, peptide-MHC complex, T cell stimulation, adoptive cell transfer, cancer immunotherapy, pharmacokinetics
- MeSH terms
- Animals, Antigens, Neoplasm, Cells, Cultured, Dendritic Cells, Humans, Lymphocyte Activation, Male, Neoplasms, Poly A, RNA, Neoplasm, Transfection, CD4-Positive T-Lymphocytes, Immunotherapy, Adoptive, Antigen Presentation, CD8-Positive T-Lymphocytes, Coculture Techniques, Cancer Vaccines, RNA Stability, Mice
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
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