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The pre-publication history for this paper can be accessed here:
|
15315712_p31
|
15315712
|
Pre-publication history
| 1.031347 |
other
|
Other
|
[
0.013091341592371464,
0.0014214670518413186,
0.9854872226715088
] |
[
0.0015875872923061252,
0.997281551361084,
0.0006836647517047822,
0.0004471398133318871
] |
en
| 0.999998 |
Integral membrane proteins are the primary choice as targets when developing new drugs and although clearly of medical relevance, forming 20% – 30% of the gene products of most genomes, these proteins have been structurally determined in only about thirty cases . Where high levels of sequence homology exist, an unknown protein's structure and hence, the location of its membrane interactive segments, can sometimes be deduced by direct comparison to known protein structures. However, where sequence information alone is available, the identification of transmembrane α -helical structure requires a bioinformatics approach to understanding the structure / function relationships of these α -helices. A number of α -helical properties have been used as models to study transmembrane α -helices and their structure / function relationships but the most commonly used are those based on the amphiphilicity of protein α -helices with the hydrophobic moment used as a measure of amphiphilicity .
|
15312230_p0
|
15312230
|
Background
| 4.172294 |
biomedical
|
Study
|
[
0.9994900226593018,
0.00022874167189002037,
0.00028117361944168806
] |
[
0.933789074420929,
0.002064957283437252,
0.06390837579965591,
0.00023763318313285708
] |
en
| 0.999994 |
To quantify the amphiphilicity of protein secondary structures, Eisenberg and co-workers introduced the hydrophobic moment, μ ( θ ), which provides a measure of the structured partitioning of hydrophilic and hydrophobic residues in a regular repeat structure of period θ . For a structure comprising L consecutive residues, the general form of μ ( θ ) is given by:
|
15312230_p1
|
15312230
|
Background
| 4.191966 |
biomedical
|
Study
|
[
0.9995792508125305,
0.0001422705827280879,
0.0002784492389764637
] |
[
0.9967179894447327,
0.001922881929203868,
0.0012795114889740944,
0.00007960225775605068
] |
en
| 0.999996 |
where H j is the hydrophobicity of the j th residue within the sequence, and θ is the angular frequency of the amino acid residues forming the structure. Eisenberg et al ., assumed that for an α -helix, θ is fixed at 100°, and that a segment of eleven consecutive residues, equivalent to three turns of an α -helix, could be used to represent amphiphilic α -helices. These assumptions led to the more generally used measure of α -helix amphiphilicity, the mean hydrophobic moment < μH >, where
|
15312230_p2
|
15312230
|
Background
| 4.242847 |
biomedical
|
Study
|
[
0.9995352029800415,
0.00015521726163569838,
0.00030950471409596503
] |
[
0.9968758821487427,
0.0012501233723014593,
0.0017837639898061752,
0.00009022329322760925
] |
en
| 0.999996 |
< μH > = μ(100°)/11
|
15312230_p3
|
15312230
|
Background
| 1.843844 |
biomedical
|
Other
|
[
0.855272114276886,
0.003158247796818614,
0.14156965911388397
] |
[
0.10976164042949677,
0.886640727519989,
0.002051853109151125,
0.0015457780100405216
] |
el
| 1 |
As a major extension to the use of the hydrophobic moment, Eisenberg et al ., introduced hydrophobic moment plot methodology, which provides a graphical technique for the general classification of protein α -helices. Using this methodology, a putative protein α -helix is characterised according to its maximum < μH > and corresponding mean hydrophobicity, < H >, where this is defined by:
|
15312230_p4
|
15312230
|
Background
| 4.119837 |
biomedical
|
Study
|
[
0.9996591806411743,
0.00011557814286788926,
0.00022525193344336003
] |
[
0.9929450154304504,
0.00335400621406734,
0.003588878083974123,
0.00011210623779334128
] |
en
| 0.999997 |
The parameters < μH > and < H > are then plotted on the hydrophobic moment plot diagram and the location of the resulting data point used to classify the putative α -helix.
|
15312230_p5
|
15312230
|
Background
| 3.275173 |
biomedical
|
Study
|
[
0.9930288195610046,
0.0006502868491224945,
0.006320917513221502
] |
[
0.8425145149230957,
0.15536370873451233,
0.001483624568209052,
0.00063812400912866
] |
en
| 0.999997 |
The mean hydrophobic moment is widely used and generally regarded as a good predictor of α -helix amphiphilicity but the results of statistical analyses have shown the efficacy of hydrophobic moment plot methodology as a predictor of α -helical class to be less certain . A number of authors have observed that the methodology can erroneously classify α -helices in cases where the hydrophobic moment for a particular amino acid sequence is greatly affected by the spatial arrangement of a few extreme amino acids, thus masking the overall nature of an α -helix . However, a more fundamental source of erroneous classification could come from the questionable assumption made by hydrophobic moment methodology with respect to angular periodicity. It is known that in naturally occurring α -helices, θ can vary over the range (95° ≤ θ ≤ 105°) and between consecutive residues . Clearly, assuming a fixed value of θ = 100° for all α -helices is an approximation and could lead to classification difficulties for the methodology. Furthermore, classification difficulties could arise from the arbitrary choice of window length made by the methodology as window length is known to have a profound effect on the relationship between < μH > and < H >. It would seem that the optimisation of θ and window length are crucial to the classification of amphiphilic α -helices yet the values chosen for these parameters by hydrophobic moment plot analysis are not optimal for the classification of any single subclass.
|
15312230_p6
|
15312230
|
Background
| 4.456615 |
biomedical
|
Study
|
[
0.998980700969696,
0.0003682996321003884,
0.000650910718832165
] |
[
0.9804107546806335,
0.0007652996573597193,
0.018667517229914665,
0.00015645683743059635
] |
en
| 0.999998 |
A number of studies have considered the significance of < μH > in relation to structure / function relationships of the α -helical classes described by hydrophobic moment plot methodology with common examples including: surface active α -helices, transmembrane α -helices and oblique orientated α -helices . However, if different α -helical classes have differing optima for θ and window length, not only does this question the validity of results obtained in these studies but also questions the validity of α -helix classification according to hydrophobic moment plot methodology. In this paper we examine the criteria upon which the methodology is based and, in view of their medical relevance, we use transmembrane α -helices as a test data set. These α -helices possess central regions, which are predominantly formed by hydrophobic residues and interact with the membrane lipid core, and end regions, which are primarily formed by hydrophilic residues and reside in the membrane surface regions . For the α -helices of our data set, we analyse the relationships for the mean hydrophobic moment and window size, angular frequency and the robustness to varying angular frequency. Correlations between the mean hydrophobic moment and mean hydrophobicity of transmembrane α -helices are established, verified and analysed to appraise biological function using resampling Bootstrap and Monte Carlo techniques .
|
15312230_p7
|
15312230
|
Background
| 4.241157 |
biomedical
|
Study
|
[
0.9992936849594116,
0.00031383128953166306,
0.0003925172204617411
] |
[
0.9990629553794861,
0.00014115376689005643,
0.0007327331113629043,
0.00006325629510683939
] |
en
| 0.999998 |
A data set of 84 transmembrane proteins were identified within Swiss-Prot and used to generate a set of 403 transmembrane sequences (see Additional file 1 ). All sequences within the data were of 21 residues in length and showed less than 5% homology (data not shown). For the sequences of this data set, the maximum mean hydrophobic moment, < μH >, and its corresponding mean hydrophobicity, < H >, were determined and used to generate the hydrophobic moment plot shown in figure 1 , based on the generally used 11 residue window ( L = 11) introduced by Eisenberg et al ., . It can be seen that data points representing the sequences of our data set cluster around the transmembrane region identified by Eisenberg et al ., but as previously noted there are a significant number that fall outside the boundaries of this region. In particular, many of this number possess < H > values less than 0.5 and would not be classified as transmembrane α -helices according to the hydrophobic moment plot taxonomy of Eisenberg et al ., . Even allowing for the diffuse nature of these boundaries on the hydrophobic moment plot diagram , these results clearly question the efficacy of hydrophobic moment methodology for the prediction of transmembrane α -helices.
|
15312230_p8
|
15312230
|
Results
| 4.207784 |
biomedical
|
Study
|
[
0.9994421601295471,
0.00024050642969086766,
0.00031728929025121033
] |
[
0.9993759989738464,
0.00016690458869561553,
0.00040618490311317146,
0.00005097589746583253
] |
en
| 0.999997 |
The above analysis was repeated except that window sizes varying in the range (7 ≤ L ≤ 20) were employed. The values for < μH > and corresponding < H > were plotted as above and the results for window sizes of 7, 9, 16 and 20 are shown in figure 2 . It can be seen that a weak negative correlation exists between < μH > and < H > for smaller window sizes but that the level of correlation appears to reduce as window size increases. The sample correlation coefficients for the various window sizes are given in table 1 . To conduct standard statistical tests to determine whether the population correlation coefficients do differ from zero, it is necessary to establish if these data are bivariate Normal. The P-values obtained from Anderson-Darling and Kolmogorov-Smirnov tests for Normality for the various window sizes with θ = 100° are shown in table 2 . These results present clear evidence that the populations for the variates for each window size are not bivariate Normal. These findings prompted the use of the bootstrap procedures to estimate the confidence intervals for the population correlation coefficient values for the window sizes in the range (7 ≤ L ≤ 20).
|
15312230_p9
|
15312230
|
Results
| 4.102739 |
biomedical
|
Study
|
[
0.9929887652397156,
0.0003779359976761043,
0.006633328273892403
] |
[
0.9996337890625,
0.00016522814985364676,
0.0001671091595198959,
0.000033947115298360586
] |
en
| 0.999996 |
The results of this investigation for θ = 100° are presented in figure 3 . It would appear that the smaller window sizes do show correlations between < μH > and < H > and if this reflects a biological property of transmembrane sequences, it could be of use in the analysis and prediction of these motifs. It is known that angular frequency for a transmembrane α -helix varies between 95° and 107° , rather than being fixed at 100° as proposed by the methodology of Eisenberg et al ., . For each window size in the range (7 ≤ L ≤ 21) residues, to accommodate the findings of Cornette et al ., , the fixed value of θ was therefore varied from 95° to 108° in increments of 1°. Once the optimal window had been obtained, to observe the discriminating effect of θ on < μH >, the < μH > values, denoted by Σ< μH >, were summed for the 403 sequences for each θ . Figure 4 shows the optimal θ , based on the maximum values of Σ< μH > for each window length. It can be seen that as the window size increases the total < μH > reduces approximately linearly until the intermediate size of eleven residues in length. For subsequent larger window sizes, we observe a further near linear reduction trend but at a reduced rate. The optimal angular frequency corresponding to each window size (7 ≤ L ≤ 21) is also given in figure 5 . The overall relationship between Σ< μH >, the window size, L , and the angular frequency, θ , is finally depicted in figure 6 as a response surface diagram.
|
15312230_p10
|
15312230
|
Results
| 4.170858 |
biomedical
|
Study
|
[
0.99918133020401,
0.00030950314248912036,
0.0005092343199066818
] |
[
0.999530553817749,
0.00012198025069665164,
0.000297915015835315,
0.00004958464342053048
] |
en
| 0.999996 |
To assess the robustness of < μH > to this fixed angular frequency assumption, and thus, the accuracy of the hydrophobic moment plot analysis for candidate transmembrane sequences, Monte Carlo simulation studies were conducted. Initially, the angular frequency, θ , was assumed to have a mean value, E( θ ), fixed at 100° and the angle for each successive residue varied about E( θ ). The random variation, X, followed a Normal distribution and six separate simulations were undertaken with X~N(100, σ 2 ), where the standard deviation, σ , was set at 0.1°, 0.3°, 0.5°, 0.7°, 0.9° and 1.1° respectively for each. The process was repeated with the mean value being set at the identified optimal angular frequency for the window size, again, for each of the window sizes in the range (7 ≤ L ≤ 20).
|
15312230_p11
|
15312230
|
Results
| 4.130951 |
biomedical
|
Study
|
[
0.9994420409202576,
0.00025516701862215996,
0.00030275926110334694
] |
[
0.9994729161262512,
0.0001615685469005257,
0.00031043903436511755,
0.00005506070010596886
] |
en
| 0.999999 |
Hydrophobic moment plots for variable angular frequency were obtained for E( θ ) = 100° for each window size in the range (7 ≤ L ≤ 21) residues and for the separate standard deviation values, σ = 0.1°, 0.3°, 0.5°, 0.7°, 0.9°, 1.1°. These were compared visually with the original plots obtained under the fixed angular frequency assumption ( θ = 100°). In all cases, the bulk properties of the plots were similar irrespective of the level of dispersion introduced by the different values of the standard deviation. The hydrophobic moment plot for L = 15; θ = 100° is provided in figure 7 . This is to be contrasted with the plots for L = 15; E( θ ) = 100°, σ = 0.1°, σ = 0.7° and σ = 1.1°, also present in figure 7 . Similar results were obtained for all other values, confirming, at least visually, that < μH > is robust to slight random perturbations about a fixed value. These properties were also observed for the simulation study with the fixed angular frequency assumption being violated about the optimum frequency for each of the window sizes in the range (7 ≤ L ≤ 20) and for each corresponding level of dispersion.
|
15312230_p12
|
15312230
|
Results
| 4.154473 |
biomedical
|
Study
|
[
0.9979844093322754,
0.00035361983464099467,
0.0016619920497760177
] |
[
0.9996032118797302,
0.00012596823216881603,
0.00023364006483461708,
0.00003721252505783923
] |
en
| 0.999997 |
A more rigorous assessment of the variation was provided by analysis of the sample correlations. These were calculated in each case and compared to the empirically derived 99% confidence intervals established for window sizes in the range (7 ≤ L ≤ 20) under the fixed angular frequency assumption of θ = 100°. The calculated sample correlation coefficients were also compared to the point estimates for the original data. In all cases, the values were within the appropriate confidence intervals and were always close to the original sample correlation coefficient values, again providing evidence that < μH > is robust to random variation in angular frequency. The results of this investigation are given in table 3 .
|
15312230_p13
|
15312230
|
Results
| 4.0697 |
biomedical
|
Study
|
[
0.9948593378067017,
0.0003595635644160211,
0.004781058989465237
] |
[
0.9996809959411621,
0.00015479209832847118,
0.00013386855425778776,
0.00003035977715626359
] |
en
| 0.999998 |
To test whether these correlations are artefactual, hydrophobic moment plots were obtained for the < μH > and < H > derived from the 403 artificial randomisation sequences generated by random re-ordering or randomisation of each of the original optimum window sequences. The plot for a window size of L = 11 is given in figure 8 . These analyses were undertaken for all those window sizes with previously identified statistically significant correlation coefficients between < μH > and < H > and were designed to test the importance of the spatial arrangement of the amino acids within the optimum sequences.
|
15312230_p14
|
15312230
|
Results
| 4.062784 |
biomedical
|
Study
|
[
0.9993022680282593,
0.00023402145598083735,
0.00046373275108635426
] |
[
0.9995941519737244,
0.00015530845848843455,
0.00020619301358237863,
0.00004441428245627321
] |
en
| 0.999997 |
Similar plots were obtained from Monte Carlo simulated data derived from the 403 sequences that had been generated by random sampling using the relative abundancies of residues found in the set of optimal windows. These analyses were therefore designed to look at the importance of relative amino acid composition for the correlations between < μH > and < H > and the results can be seen for a window size of L = 11 in figure 8 . Again, analyses were performed for all window sizes with associated statistically significant correlations (data not shown). It is worth noting that since the effect of varying window size had a significant effect on the correlation between < μH > and < H >, varying L was observed to vary the optimal sequence identified within the transmembrane domain. Clearly this was not unexpected.
|
15312230_p15
|
15312230
|
Results
| 4.084045 |
biomedical
|
Study
|
[
0.999327540397644,
0.0002524364972487092,
0.00042009545722976327
] |
[
0.9995713829994202,
0.0001542877871543169,
0.00022461608750745654,
0.000049715985369402915
] |
en
| 0.999996 |
It can be seen from figure 5 that the most discriminating angular frequency for a fixed window size varies within the range, (95° ≤ θ ≤ 104°) for window sizes (7 ≤ L ≤ 20). There is an obvious damped oscillation present, which can be seen to correspond to the assumed intrinsic periodicity of α -helical secondary structure i.e. 3.6 residues per turn. Figure 5 clearly demonstrates that the fixed 100° angular frequency, assumed when modelling α -helices in general, is no more than a representative average with a value nearer 102° providing a maximum for an optimum L = 11 residue window in a transmembrane α -helical sequence.
|
15312230_p16
|
15312230
|
Conclusions
| 4.17994 |
biomedical
|
Study
|
[
0.9989213943481445,
0.0002610302472021431,
0.0008175888215191662
] |
[
0.9994217157363892,
0.0002832978789228946,
0.00025165241095237434,
0.00004342910324339755
] |
en
| 0.999997 |
From figure 4 , it is also evident that the degree of discrimination possible using < μH > declines in a near linear fashion with increasing window size with the optimum L = 11 residue window appearing to provide approximately average discrimination for transmembrane α -helices. The bootstrap derived 99% confidence intervals for the correlation coefficients between < μH > and < H > for window sizes in the range (7 ≤ L ≤ 20) showed that there are highly significant linear associations for the smaller window sizes in the range (7 ≤ L ≤ 16). As the magnitude of each of the corresponding sample coefficients is small (table 1 ), this should be interpreted as evidence of a strong (negative) association but with high variability being present. These correlations become weaker, on average, with increasing window size until they are not statistically significant at the 1% level and we have no compelling evidence that the variates are not independent. The choice of window size therefore, becomes paramount if < H > and < μH > are to be used to classify transmembrane α -helices. More importantly, the variation in correlation between these parameters and the effect of varying window size on the location of the sequence identified as optimal for α -helix classification brings into question the relevance of using the mean hydrophobic moment for comparison between varying window sizes. However, < μH > has been shown to be robust to departures from the fixed angular frequency assumption for a large range of window sizes appropriate for transmembrane proteins and for a range for levels of dispersion.
|
15312230_p17
|
15312230
|
Conclusions
| 4.263573 |
biomedical
|
Study
|
[
0.9993413090705872,
0.00032517878571525216,
0.00033348725992254913
] |
[
0.999049961566925,
0.0001946283009601757,
0.0006830708589404821,
0.00007237787212943658
] |
en
| 0.999997 |
There were no substantial differences between the plots for relative abundance sample data and those for the randomisation sequences except for a few chance negative < H > observations from the former. This suggests that there are no serial correlations between residue types, where presence in the identified section of the penetrating transmembrane stretch is determined predominantly by relative abundance. This is to be contrasted with the distribution of observations for the original transmembrane sequences for a window size of 11 residues . Most noticeable is the difference in < μH > over the range of < H > values. There appears to be a lower bound for < μH > for the original sequence, which is clearly not present for the randomisation data. Furthermore, whilst the negative correlation would appear to be an artefact, as it is exhibited in all cases, the dispersion around any optimal fitted line through the data such as a least squares fit also is clearly different. It appears similar and quite spread out for the two randomised sequence data but considerably less so for the transmembrane sequences. This provides evidence that within the optimum window, whilst residue composition is not influential, order is. It would appear that this ordering is leading to both organisation and biological function for at least segments of the interacting portions of transmembrane proteins. This is consistent with the belief that the hydrophobic moment is a good predictor of amphiphilicity although it can be unduly influenced by relatively few amino acid residues within a sequence .
|
15312230_p18
|
15312230
|
Conclusions
| 4.32516 |
biomedical
|
Study
|
[
0.999288022518158,
0.0003907131322193891,
0.0003212711599189788
] |
[
0.999040424823761,
0.0002344439271837473,
0.0006401014397852123,
0.00008494641224388033
] |
en
| 0.999997 |
In summary, our analyses confirm previous studies, which have shown limitations to the ability of hydrophobic moment plot methodology to assign function to membrane interactive α -helices . More importantly, our investigation leads to a questioning of the logic of comparing mean hydrophobic moments, in general, for transmembrane proteins. This is due to the effect of window size on both, the correlation of mean hydrophobic moment with mean hydrophobicity and the identified sensitivity of the optimum window. Comparisons of the hydrophobic moment are seemingly only meaningful for separate transmembrane proteins with identical window sizes.
|
15312230_p19
|
15312230
|
Conclusions
| 4.127457 |
biomedical
|
Study
|
[
0.9994871616363525,
0.0002480482216924429,
0.0002647176443133503
] |
[
0.9987913966178894,
0.00017554208170622587,
0.0009688499849289656,
0.00006427615153370425
] |
en
| 0.999995 |
Despite these limitations, < μH > has been shown to be robust to departures from the fixed angular frequency assumption for transmembrane proteins. Given the severe lack of structural information for transmembrane proteins, the identification of transmembrane α -helices using hydrophobic moment based analyses, and other bioinformatic approaches, seems likely to continue for the foreseeable future. Nonetheless, the results of such analyses should only be taken as a guide, and where possible, obtaining corroborative experimental data is essential. On the positive side, our results have demonstrated the importance of amino acid residue sequence order in establishing organisation and biological function for the transmembrane α -helices of proteins. With the ongoing development of predictive techniques, these results should be useful in furthering this development and helping to improve drug target identification.
|
15312230_p20
|
15312230
|
Conclusions
| 4.121413 |
biomedical
|
Study
|
[
0.9996337890625,
0.00018670735880732536,
0.00017942872364073992
] |
[
0.9962496161460876,
0.0003771711199078709,
0.003284834325313568,
0.00008834913751343265
] |
en
| 0.999995 |
The primary structures of 96 transmembrane proteins were selected from the Swiss-Prot data bank ( ; accessed 25.05.04) and confirmed as transmembrane by extensive analysis of the literature. The sequences were analysed for homology using the sequence alignment program BLAST (Basic local alignment search tool) and twelve homologous sequences were rejected. From the remaining 84 primary structures, a data set comprising 403 putative transmembrane α -helical sequences, each of 21 residues, was established using the algorithm, Top Pred2 (; ; accessed 25.05.04).
|
15312230_p21
|
15312230
|
The selection of transmembrane, α -helix forming segments
| 4.151532 |
biomedical
|
Study
|
[
0.9995518326759338,
0.0001865778031060472,
0.00026159986737184227
] |
[
0.9991787075996399,
0.00026177268591709435,
0.0005067025194875896,
0.00005276671799947508
] |
en
| 0.999996 |
In the present study, all hydrophobic moment plot analyses were performed using the consensus hydrophobic scale of Eisenberg . To identify putative transmembrane α -helix forming segments using hydrophobic moment plot methodology, hydropathy plot analysis is initially undertaken to identify the primary amphiphilicity of candidate sequences. These sequences are selected using a 21 residue window as this is sufficiently long for an α -helix to span the bilayer.
|
15312230_p22
|
15312230
|
Hydrophobic moment plot analysis of transmembrane, α -helix forming segments
| 4.106132 |
biomedical
|
Study
|
[
0.9993745684623718,
0.00020255935669410974,
0.00042287996620871127
] |
[
0.9995673298835754,
0.00024078969727270305,
0.00015083920152392238,
0.000041046965634450316
] |
en
| 0.999998 |
Once a putative transmembrane domain has been identified, an eleven residue window is considered to progress along the amino acid sequence and for each window, the hydrophobic moment at 100° is calculated. Based on the assumption that a protein sequence will adopt its most amphiphilic arrangement, the window with the maximum mean hydrophobic moment, < μH >, is taken as the most likely to form an amphiphilic α -helix . The location of the optimum window was observed accordingly for window sizes of seven through to twenty consecutive residues.
|
15312230_p23
|
15312230
|
Hydrophobic moment plot analysis of transmembrane, α -helix forming segments
| 4.189505 |
biomedical
|
Study
|
[
0.999535083770752,
0.00023441374651156366,
0.00023054170014802366
] |
[
0.9989269375801086,
0.0006569754332304001,
0.00034256032085977495,
0.00007343264587689191
] |
en
| 0.999997 |
For window sizes ranging from 7 to 20 amino acid residues < μ H > were computed for the range of angular frequency values (95° ≤ θ ≤ 108°). In each case, the value of θ , which maximises < μ H >, i.e. the value of θ which produces < μH >, was determined and is referred to as the optimal angular frequency for that window size. These procedures were based on previously published work, which identified variations in θ for α -helices .
|
15312230_p24
|
15312230
|
Optimal angular frequency and window length for < μH >
| 4.088031 |
biomedical
|
Study
|
[
0.9993744492530823,
0.0002128638152498752,
0.0004127202555537224
] |
[
0.9992437362670898,
0.00034536770544946194,
0.0003555827133823186,
0.00005540706479223445
] |
en
| 0.999998 |
For window sizes ranging from 7 to 20 amino acid residues, scatterplots of < μH > versus < H > (hydrophobic moment plots) with θ = 100° were obtained. The corresponding sample correlation coefficients were calculated to identify the effect of window size on the relationship between these variates and hence on their ability to act as discriminators in the prediction of transmembrane α -helices. In addition, for each window size in the range (7 ≤ L ≤ 20) residues and for θ in the range (95° ≤ θ ≤ 108°), the response surface diagram for < μH > was constructed.
|
15312230_p25
|
15312230
|
Hydrophobic Correlation
| 4.125694 |
biomedical
|
Study
|
[
0.9994685053825378,
0.0002537054824642837,
0.0002778208290692419
] |
[
0.9992642998695374,
0.00018640514463186264,
0.0004884506925009191,
0.000060936530644539744
] |
en
| 0.999997 |
Statistical confidence intervals were established for the Pearson (Product-Moment) Correlation Coefficient between < μH > and < H > for both cases where window size was varied for a fixed value of the angular frequency, and the angular frequency was varied for a fixed window size. The resulting mean hydrophobicity measures were checked for bivariate Normality and non-parametric bootstrap procedures were used to estimate confidence intervals for the Correlation Coefficients .
|
15312230_p26
|
15312230
|
Confidence intervals for the Correlation Coefficient
| 4.08737 |
biomedical
|
Study
|
[
0.9991410970687866,
0.00021728135470766574,
0.0006416551768779755
] |
[
0.9994760155677795,
0.0002354502066737041,
0.0002486318408045918,
0.00003994749567937106
] |
en
| 0.999997 |
To provide evidence of the statistical significance of any linear association, the bootstrap bias-corrected and accelerated technique (BCa) and an analytical extension of this, the ABC . In addition, the bootstrap Delta method was employed, which although another bootstrap based method, was developed specifically for estimating the variance of a function of sample means. As the sample Correlation Coefficient can be readily expressed as such a statistic, it is also well suited to the estimation of confidence intervals for these Correlation Coefficients . As both main approaches differ substantially, a more informed assessment of statistical significance could therefore be made.
|
15312230_p27
|
15312230
|
Confidence intervals for the Correlation Coefficient
| 4.06439 |
biomedical
|
Study
|
[
0.9974731802940369,
0.00020968748140148818,
0.0023171803914010525
] |
[
0.9969832301139832,
0.0024940131697803736,
0.0004687891050707549,
0.000053972387831890956
] |
en
| 0.999998 |
To assess the robustness of < μH > to the fix angular frequency assumption, e.g., θ = 100°, θ was varied randomly about 100° and < μH > was calculated for each of the optimal windows for window sizes (7 ≤ L ≤ 20) for the 403 transmembrane proteins. These calculations were also obtained for similar random variations about the observed optimum angular frequencies, again, for the various window sizes (7 ≤ L ≤ 20). In all cases, it is assumed that the variation follows a Normal distribution with the mean value set at the desired value for θ and with the standard deviation, σ , set at: 0.1°, 0.3°, 0.5°, 0.7°, 0.9° and 1.1° respectively for six separate Monte Carlo simulation studies. The sample correlation coefficients for each simulation were calculated and compared to the empirically derived 99% confidence intervals for the corresponding population values and, in particular, with the point estimates for the original sequences.
|
15312230_p28
|
15312230
|
Variable angular frequencies
| 4.10836 |
biomedical
|
Study
|
[
0.9993435740470886,
0.00025797050329856575,
0.00039851857582107186
] |
[
0.9995042085647583,
0.00014153227675706148,
0.0003050855593755841,
0.00004910270217806101
] |
en
| 0.999997 |
Given that these data are from an observational study, it is necessary to assess whether any linear associations between < μH > and < H > for the α -helix forming sequences of our data set are likely to be causal or merely an artefact of amino acid composition. To investigate these possibilities, two additional simulation studies were undertaken. The first looked at spatial arrangements of residues within the primary sequences and the second focused on the effect of amino acid composition on correlations between < μH > and < H >.
|
15312230_p29
|
15312230
|
Causality and biological function
| 4.019601 |
biomedical
|
Study
|
[
0.9992451667785645,
0.00023506111756432801,
0.0005197844002395868
] |
[
0.999543309211731,
0.00020631254301406443,
0.00019974065071437508,
0.000050692364311544225
] |
en
| 0.999996 |
To assess if positional or sequential correlational properties existed for the amino acids within the sequences, the sequence of residues for each of the optimum windows was re-ordered randomly. Artificial sequences were thus generated by random rearrangement or randomisation of the primary sequences within the 403 optimal windows. Hence, each window associated with < μH > was used to generate a random arrangement.
|
15312230_p30
|
15312230
|
Causality and biological function
| 4.037595 |
biomedical
|
Study
|
[
0.9994663596153259,
0.00021289658616296947,
0.0003207357367500663
] |
[
0.9993075132369995,
0.00034667481668293476,
0.00028914347058162093,
0.00005664007767336443
] |
en
| 0.999997 |
To further investigate whether correlations between < μH > and < H > were dependent on sequence composition and not on spatial or sequential correlation, an additional parametric bootstrap simulation study was conducted. Here 403 artificial sequences were created. Each was randomly generated where, for each position, selection was based on the relative abundance of all the residues for the complete 403 optimum windows.
|
15312230_p31
|
15312230
|
Causality and biological function
| 4.020144 |
biomedical
|
Study
|
[
0.9989558458328247,
0.0002614112163428217,
0.0007827094523236156
] |
[
0.9996408224105835,
0.00017080492398235947,
0.00014678375737275928,
0.00004153466215939261
] |
en
| 0.999995 |
In both cases the corresponding < μH > and < H > from these newly created sequences were calculated, the associated hydrophobic moment plots obtained and sample correlations calculated. These were inspected to assess whether any linear associations for the original transmembrane data were thus likely to be causal or merely artefactual and whether, from inspection of variation, there was evidence of increased organisation, which could be interpreted as an indication of biological function.
|
15312230_p32
|
15312230
|
Causality and biological function
| 4.011152 |
biomedical
|
Study
|
[
0.9993801116943359,
0.0002763200318440795,
0.0003435506660025567
] |
[
0.9994555115699768,
0.00023962429258972406,
0.0002468811289872974,
0.000058023157180286944
] |
en
| 0.999996 |
The dimeric feature of the retroviral RNA genome was identified almost forty years ago. However, as with many topics in retrovirology, interest in this area was heightened with the realization that the causative agent of AIDS was a retrovirus. Since then, RNA and protein sequences involved in genome dimerization have been identified for a number of retroviruses, and the dimeric nature of the retroviral genome is known to be important for various critical events in the viral life cycle. These include reverse transcription and recombination, as well as genome encapsidation. To date, a number of informative reviews have been published on retroviral RNA dimerization , genome packaging , and the role of nucleocapsid (NC) protein in these activities . More recently, a comprehensive review was published that summarized the contributions of in vitro analysis to the identification of retroviral dimerization signals, and provided an overview of the HIV-1 5' untranslated region (UTR) structure with reference to a number of proposed models . Another, in this issue of Retrovirology , focuses on the different roles of different dimer linkage structures amongst various retroviruses . In this review, we will focus on results from in vivo studies that provide insights into the relationship between retroviral RNA dimerization and packaging, and the biological relevance of these activities to viral replication.
|
15345057_p0
|
15345057
|
Introduction
| 4.268687 |
biomedical
|
Review
|
[
0.978576123714447,
0.012590655125677586,
0.00883321464061737
] |
[
0.003780802246183157,
0.0020857034251093864,
0.9931052327156067,
0.0010282128350809216
] |
en
| 0.999996 |
The first evidence for the existence of a dimerized RNA genome came in 1967 when it was shown that viral RNA from each of Rous sarcoma virus (RSV), avian myeloblastosis virus (AMV), murine leukemia virus (MLV), and mouse mammary tumor virus (MTV) displayed sedimentation constants between 64S and 74S in sucrose gradients . Since these sedimentation constants and corresponding molecular weights were much larger than those of most other known viral RNAs, the structure of these RNA genomes became a matter of great interest. Experiments showing that the 62S RSV RNA species could be converted to a 36S species by heat treatment suggested a disaggregation of the 62S RNA into smaller RNAs, and implied that the fast-sedimenting (62S) RSV RNA was actually an aggregate of smaller (36S) RNAs . The first real understanding of this putative aggregate RNA structure came in 1975 when RNA from the endogenous feline retrovirus, RD-114, was visualized by electron microscopy (EM), and it was apparent that the 52S RNA molecule existed as an extended single strand that contained a central Y- or T-shaped secondary structure . It appeared that this 52S molecule actually consisted of two half-size molecules, joined together by the Y- or T-shaped structure, which was termed rabbit ears (RE). It was later shown that the RNA had a poly(A) sequence at each of the two free ends. More importantly, this indicated that nucleotides involved in this RE, or dimer linkage structure (DLS), resided in the 5' region of the RNA genome . Similar structures were also reported for numerous other type C RNA viruses . The absolute conservation of a DLS among retroviruses was strong evidence that the dimerization process must be critical to the retroviral life cycle. With the discovery that the causative agent of AIDS was also a retrovirus, inhibition of RNA dimerization was proposed as a possible therapy for HIV, and HIV-1 RNA dimerization became an intensely studied topic.
|
15345057_p1
|
15345057
|
Retroviral RNA dimerization
| 4.871916 |
biomedical
|
Review
|
[
0.9958159327507019,
0.0024064513854682446,
0.0017775249434635043
] |
[
0.3802928030490875,
0.0034572449512779713,
0.6138264536857605,
0.002423530677333474
] |
en
| 0.999997 |
Both in vivo and in vitro approaches have been used to study retroviral RNA dimerization. The in vivo approach is that whereby RNA is isolated from virions produced in tissue culture and then analyzed by native Northern blotting . The other method involves synthesis of short segments of viral RNA in vitro , and then studying the ability of these fragments to form dimers. The HIV-1 DLS was originally identified when it was shown that an in vitro -transcribed fragment of HIV-1 RNA could form two major bands on a native gel after incubation at 37°C for 15 min . The lower band had the expected size of the RNA fragment, while the upper band corresponded to a dimer. In vivo evidence for a role of the NC protein in the dimerization process was already available , and this study also showed that NC could bind to viral RNA and increase the rate of dimerization of the RNA fragments in these in vitro dimerization assays .
|
15345057_p2
|
15345057
|
Retroviral RNA dimerization
| 4.222404 |
biomedical
|
Study
|
[
0.9996078610420227,
0.00023147255706135184,
0.00016073841834440827
] |
[
0.9986636638641357,
0.00026347438688389957,
0.000994157511740923,
0.00007865310180932283
] |
en
| 0.999998 |
It was subsequently reported that an RNA fragment representing nt 1–311 of HIV-1 RNA (Mal strain; a chimera of subtypes A and D) could not only form dimers, but that RNAs containing these first 311 nt could dimerize 10 times faster than RNA sequences at positions 311–415 that were previously shown to be sufficient for HIV-1 RNA dimerization . Based on these results, the authors concluded that sequences upstream of the splice donor site are involved in the dimerization process, and proposed that sequences in this region somehow hastened the reaction. The key nucleotides involved in this RNA dimerization event make up a palindromic sequence, 274-GUGCAC-279, between the PBS and the major splice donor , and RNA sequences on both sides of this palindrome can form a stem-loop structure with the palindrome in the hairpin loop. Deletion of this stem-loop motif (nt 265–287) completely abolished dimerization of the 1–615 HIV-1 RNA fragment in vitro . The palindromic region was termed the dimerization initiation site (DIS) and it was proposed that this structural element could be exploited for targeted antiviral therapy by antisense oligonucleotides . These findings were later confirmed when a 19 nt sequence upstream of the 5' major SD was shown to be part of the HIV-1 RNA dimerization domain (Lai strain; subtype B) , and it was found that in vitro dimerization of a 224–402 nt RNA fragment was completely blocked by an antisense oligonucleotide that targeted the palindrome . This led to a "loop-loop kissing complex" or "kissing-loop model" of HIV-1 RNA dimerization, in which the 6 nt palindromes on each of the two monomeric RNA molecules interact through Watson-Crick base-pairing. Purine residues flanking the palindrome were later shown to be intricately involved in this initial interaction which is believed to shift the equilibrium toward the formation of dimers, allowing the stems to melt and anneal to their complementary sequences on the other RNA molecule, thus forming the stable extended duplex . This model fits with the idea that immature virions contain a less stable dimer involving only base-pairing of the palindromes, but that the mature virions contain a more stable structure, the extended duplex. Subsequent phylogenetic analysis of over 50 HIV-1, HIV-2, and simian immunodeficiency virus (SIV) nucleotide sequences showed an absolute conservation of a predictable structure similar to the DIS, with the hallmark of the HIV-1 DIS motif being a 6 nt palindrome consisting of either a GCGCGC or a GUGCAC sequence . Similar kissing-loop models have also been proposed for a number of other retroviruses .
|
15345057_p3
|
15345057
|
Retroviral RNA dimerization
| 4.922299 |
biomedical
|
Study
|
[
0.9977070093154907,
0.001657123677432537,
0.0006358519312925637
] |
[
0.9735034108161926,
0.001815886702388525,
0.023303287103772163,
0.0013773387763649225
] |
en
| 0.999998 |
Despite ample in vitro evidence supporting the above model of dimer maturation, it was not yet known where or when the RNA dimer was actually formed in vivo . However, native Northern blotting analysis of RNA from two Moloney murine leukemia virus (MuLV) protease-negative (PR - ) mutants displayed dimers that migrated more slowly, and showed lower melting temperatures, than that of wild-type . It was therefore concluded that PR function is required for RNA maturation in MuLV. Similar experiments with a related virus also suggested that the RNA maturation event required an intact, unsubstituted Cys array within the NC domain . On the basis of these results, a maturation pathway was proposed for MuLV in which Gag polyprotein molecules assemble into a nascent virion containing an immature dimer. The particle would then be released from the cell, and once Gag is cleaved by PR, NC would act on the immature dimer, converting it to the mature form.
|
15345057_p4
|
15345057
|
Retroviral RNA dimerization
| 4.441244 |
biomedical
|
Study
|
[
0.9994551539421082,
0.0003481730236671865,
0.00019667331071104854
] |
[
0.9984690546989441,
0.0005344212986528873,
0.000852699507959187,
0.00014374744205269963
] |
en
| 0.999998 |
Evidence for the role of NC in this dimer maturation process came when in vitro analysis showed that NC could convert the less thermostable dimers to a more stable conformation . Similar results were obtained by others showing that HIV-1 NC could activate dimerization of a 77–402 nt fragment of HIV-1 Lai RNA, as well as convert an unstable dimer, corresponding to the kissing complex, to a stable one . Taken together, these thermostability conversions seem to resemble the RNA maturations reported in vivo , and, in agreement with earlier proposals , strongly suggest that NC is responsible for the dimer maturation depicted in Fig. 1 . Subsequent in vivo analysis of a panel of HIV-1 NC mutants showed that Cys-Ser substitution of amino acid residues within the second zinc finger decreased genomic RNA dimerization to the same extent as disruption of the DIS . This finding confirmed the involvement of NC in the dimerization process, and suggests that the kissing-loop model also applies to the in vivo situation.
|
15345057_p5
|
15345057
|
Retroviral RNA dimerization
| 4.440151 |
biomedical
|
Study
|
[
0.9994627833366394,
0.00034913429408334196,
0.00018809072207659483
] |
[
0.9983285069465637,
0.0003395674575585872,
0.0011967808241024613,
0.0001351418613921851
] |
en
| 0.999998 |
Why a class of viruses would evolve to have such a unique genomic structure is not entirely clear, but it is speculated that the availability of two copies of the genome would be advantageous for recombination during the complex reverse transcription process, that is key to the retroviral life cycle . Indeed, the dimeric nature of the genome is thought to be responsible for a high rate of recombination during infection . Given that most dimerization signals overlap with known packaging elements, it was naturally assumed that it is the RNA dimer that is specifically recognized for packaging in the case of retroviruses, and that this dimeric feature ensures proper packaging of two copies of genomic RNA. A number of studies have attempted to address this question of a link between dimerization and packaging, but let us first review several aspects of the HIV-1 RNA packaging process.
|
15345057_p6
|
15345057
|
HIV-1 RNA packaging
| 4.222495 |
biomedical
|
Review
|
[
0.998268723487854,
0.0008742362260818481,
0.0008570686331950128
] |
[
0.13663770258426666,
0.0024985505733639,
0.8601855635643005,
0.00067818199750036
] |
en
| 0.999994 |
The first studies aimed at identifying the HIV-1 RNA packaging signal found that deletion of RNA sequences between the major splice donor (SD) and the gag coding region decreased the levels of genomic RNA packaged into virions . Since these sequences were downstream of the major 5' SD, and therefore would not be found in any spliced viral RNA species, it was plausible that this region could be responsible for the selective packaging of genomic RNAs. Analysis of the putative ψ locus from a variety of retroviruses showed that these sequences had the ability to direct the selective encapsidation of heterologous RNAs to which they had been linked artificially . In HIV-1, such autonomous packaging signals were mapped to the regions extending 30–40 nt immediately upstream and downstream of the gag start codon ; however, subsequent studies showed that RNA sequences upstream of the 5' SD site also affected RNA packaging . It was also known that retroviral encapsidation required trans-acting amino acid sequences in the Gag protein , and several groups reported that HIV-1 Gag and NC exhibit specific binding affinity for the HIV-1 ψ site in vitro . These findings, combined with chemical and RNase accessibility mapping, as well as computerized sequence analysis, led to the generation of a model for the HIV-1 ψ site that comprised four independent stem-loops . Three of these hypothetical stem-loop structures were each shown to serve as independent Gag binding sites, and were proposed to contribute individually to overall packaging efficiency. SL1, SL3, and SL4 were later shown to be critical for packaging specificity in vivo . Subsequent in vitro analysis from another group demonstrated that the major packaging signal is an extended bulged stem-loop whose RNA conformation is altered upon interaction with Gag . However, more recent work indicates that SL2 and SL3 display much higher affinities for NC than SL1 and SL4 in vitro . Based on these findings, a model has been proposed to represent the initial complex formed between the NC domains of assembling Gag molecules and the dimeric ψ region . In this model, SL1 is shown to form an RNA duplex between the two stands, while SL4, instead of directly binding to Gag, contributes additional RNA-RNA interactions that stabilize the tertiary structure of the ψ element. The RNA conformation resulting from this folding pattern is thought to expose SL2 and SL3 for high-affinity binding to Gag.
|
15345057_p7
|
15345057
|
HIV-1 RNA packaging
| 4.794687 |
biomedical
|
Study
|
[
0.9983892440795898,
0.001110768411308527,
0.0005000183591619134
] |
[
0.9798163771629333,
0.0009772416669875383,
0.018570711836218834,
0.0006356446538120508
] |
en
| 0.999996 |
Despite the clear results obtained from simplified in vitro studies such as those mentioned above, the SL1-4 region alone is not sufficient to target RNA into HIV-1 virions in vivo , and the minimal region required to confer autonomous packaging activity actually maps to a larger region covering the first 350–400 nt of the genome, including ≈ 240 nt upstream of SL1 . In agreement with these studies, mutations that alter the stability of the poly(A) hairpin stem region, or delete the upper part of the hairpin, severely inhibited HIV-1 replication . And, these deficits in replication correlated with reduced RNA packaging levels in virions, suggesting that the formation of the poly(A) hairpin is necessary for normal packaging of viral genomes. Subsequent research confirmed the importance of the poly(A) hairpin in the RNA packaging process , and it was shown that similar disruption of base-pairing in the stem of the TAR element also caused profound defects in packaging . Finally, deletion analyses of RNA sequences between the poly(A) hairpin and SL1 suggested that unspecified sequences within the U5-PBS region also contribute to HIV-1 RNA packaging . Our group later showed that GU-rich sequences in the lower stems of the poly(A) hairpin and the U5-PBS complex contribute to both dimerization and packaging .
|
15345057_p8
|
15345057
|
HIV-1 RNA packaging
| 4.532924 |
biomedical
|
Study
|
[
0.9992114305496216,
0.0005373154999688268,
0.00025126137188635767
] |
[
0.9975255131721497,
0.00045803363900631666,
0.0018044919706881046,
0.00021187086531426758
] |
en
| 0.999996 |
In summary, all of the seven predicted stem-loop structures in the HIV-1 5' UTR are known to be important for genome encapsidation, and all of these RNA structural elements have also been assigned other functions in various steps of the viral life cycle, e.g. the role of SL1 in the initiation of dimerization. The existence of such overlapping functions for these RNA structures raises the possibility that some of these functions, such as dimerization and packaging, might be linked. The evidence for and against the existence of such a link in HIV-1 will be the main focus of the remainder of this review.
|
15345057_p9
|
15345057
|
HIV-1 RNA packaging
| 4.040782 |
biomedical
|
Review
|
[
0.9910535216331482,
0.004199931863695383,
0.004746559076011181
] |
[
0.004592643119394779,
0.0014877411304041743,
0.99347323179245,
0.00044640363194048405
] |
en
| 0.999998 |
One of the first electron microscopy studies of a retroviral DLS in 1976 proposed that this region "could have some role in packaging the RNA in the virus" . This raised the question of a possible link between dimerization and packaging that is still debated. The answer to this question has significance in our basic understanding of the retroviral life cycle and may also have implications for therapy, since many groups are actively studying these two activities as potential drug targets.
|
15345057_p10
|
15345057
|
Is dimerization a prerequisite for packaging?
| 3.796895 |
biomedical
|
Review
|
[
0.9989136457443237,
0.0003676509950309992,
0.000718739174772054
] |
[
0.21195824444293976,
0.06542003154754639,
0.7214749455451965,
0.0011468138545751572
] |
en
| 0.999996 |
Early reports on in vitro dimerization of HIV-1 RNA showed that the DLS localized to a stretch of genomic RNA downstream of the 5' SD (nt 311–415) , and it was noted that this dimerization domain encompassed a previously identified packaging element that had also been shown to bind NC . This dependence of HIV-1 RNA dimerization on cis elements required for packaging was immediately interpreted to mean that retroviral RNA dimerization, activated by either NC or Gag precursors, should direct genomic RNA into the virion, implying that dimerization might be a prerequisite for packaging. Since HIV-1, MuLV, and RSV all contain elements involved in dimerization that were also required for packaging , it was proposed that dimerization might function as a molecular switch that negatively regulates translation and positively regulates encapsidation . The existence of a DLS downstream of the major splice donor would seemingly supply a convenient mechanism whereby only genome length RNA would be able to dimerize and subsequently become encapsidated into the virion. However, evidence questioning such a dimerization-mediated mechanism of genomic RNA packaging came from studies showing that sequences upstream of the SD site had even greater dimerization capabilities than those located downstream . The involvement of such sequences (e.g. the DIS, SL1) in the dimerization process questioned the link between dimerization and packaging, because these sequences are also found in all HIV-1 spliced viral RNAs.
|
15345057_p11
|
15345057
|
Clues from in vitro studies
| 4.761307 |
biomedical
|
Study
|
[
0.9989513158798218,
0.0006532313418574631,
0.00039543627644889057
] |
[
0.9667484760284424,
0.001019657007418573,
0.031720105558633804,
0.0005117411492392421
] |
en
| 0.999996 |
Early in vivo studies analyzing the structure of virion-associated RNA from rapid-harvest avian retroviruses showed that viral RNA appeared to be a mixture of monomers and dimers . Similar results had also been reported with PR and NC mutants, which argued against the notion that dimerization is a prerequisite for packaging. However, analysis of rapid-harvest virus in MuLV showed that genomic RNA was already in the form of a dimer shortly after budding, albeit as a less stable, physically different RNA dimer than that present in mature virions . Based on these observations it was proposed that MuLV particles never package monomeric RNAs, but rather that the dimeric RNA structure might be integral to the packaging signal that is recognized by Gag during assembly. It was also speculated that the previously reported presence of monomers in viral RNA preparations had resulted from the physical dissociation of fragile unstable dimers during RNA preparation. Similar experiments performed on PR - mutants of HIV-1 showed that substantial amounts of monomeric RNA could be detected . Since PR - dimers were shown to be less stable than wild-type dimers, it was assumed that dimers were preferentially packaged in PR - particles, but that some fragile dimeric structures had dissociated during RNA preparation. Based on these in vivo results with both MuLV and HIV-1, it was concluded that dimerization is a prerequisite for packaging and should be considered to be a general feature of retrovirus assembly.
|
15345057_p12
|
15345057
|
Observations from in vivo studies
| 4.757186 |
biomedical
|
Study
|
[
0.9988705515861511,
0.0006357055972330272,
0.000493653875309974
] |
[
0.9676815867424011,
0.0008664241177029908,
0.03101302497088909,
0.0004389940295368433
] |
en
| 0.999997 |
Further insights into this topic can be obtained by examination of results from a number of studies aimed at understanding the role of the DIS in HIV-1 replication. One such study, in which DIS loop palindrome sequences were mutated, found that mutation of the palindrome to shorter or longer versions of GC stretches did not have major effects on viral RNA dimerization; however, partial RNA packaging defects were observed that also corresponded to diminutions in viral replication . Based on these data, it was proposed that these DIS loop mutants might have experienced a partial dimerization defect that caused inefficient packaging . In a similar study, mutation of the palindrome, as well as deletion of the upper stem-loop of SL1 caused drastic reductions in viral infectivity and decreases in both dimerization and packaging of HIV-1 genomic RNA . In an attempt to explain how these mutations could affect both activities, a model was proposed in which Gag does not specifically recognize the dimerized genome but rather initially interacts with one molecule of genomic RNA that happens to be linked (dimerized) to a second such molecule. Then, during packaging, Gag would effectively bind to two genomic RNA molecules at once. Hence, defects in dimerization would result in subsequent packaging defects. Based on these data, it was also concluded that the encapsidation and dimerization processes are coupled to some extent.
|
15345057_p13
|
15345057
|
Observations from in vivo studies
| 4.47648 |
biomedical
|
Study
|
[
0.9993466734886169,
0.0004048589034937322,
0.00024846504675224423
] |
[
0.9931926131248474,
0.00038103980477899313,
0.0062512606382369995,
0.00017509559984318912
] |
en
| 0.999996 |
Although several groups had attempted to delineate the relationship between dimerization and packaging, the fact remains that the RNA signals that are important for both of these activities overlap in most retroviral genomes; this makes it difficult to interpret the results of mutagenesis studies. In an attempt to generate viruses that would be expected to display selective defects in dimerization or packaging, one group designed a panel of constructs containing mutations in SL1, SL3, or both . Results from this study showed that deletion of either SL1 alone, or SL3 plus adjacent flanking sequences, reduced genomic packaging, while deletion of SL1 and SL3 simultaneously caused an even further reduction. With respect to dimerization, complete deletion of SL1, or even disruption of the base-pairing in the upper stem, resulted in elevated levels of monomer-sized RNA species on native Northern blots, again confirming the importance of this region for the in vivo HIV-1 RNA dimerization process. Yet, these mutant genomes could still be packaged, suggesting that HIV-1 RNAs need not be dimers for this to happen. Thus, the authors concluded that dimerization is not a prerequisite for packaging but rather serves an independent function in the retroviral life cycle. In the above-cited article, the effects of SL3 mutations on dimerization were not studied, but our group later showed that viruses containing even minor substitutions in or around SL3 could have significant effects on both dimerization and packaging .
|
15345057_p14
|
15345057
|
Observations from in vivo studies
| 4.278669 |
biomedical
|
Study
|
[
0.9994878768920898,
0.00030098759452812374,
0.00021112393005751073
] |
[
0.9984686970710754,
0.00023109371250029653,
0.0012081407476216555,
0.00009205203969031572
] |
en
| 0.999996 |
In summary, the in vivo studies described above commonly observed that mutations in 5' RNA sequences affected both dimerization and packaging, presumably due to the close proximity of the RNA dimerization and packaging signals.
|
15345057_p15
|
15345057
|
Observations from in vivo studies
| 4.000184 |
biomedical
|
Study
|
[
0.9995580315589905,
0.00019275069644208997,
0.0002491519262548536
] |
[
0.9848712086677551,
0.0013369061052799225,
0.013649199157953262,
0.0001426513190381229
] |
en
| 0.999994 |
In an attempt to separate the dimerization and packaging functions, and to characterize the DIS-DLS region without altering packaging activity, one group generated mutant constructs carrying a duplication of approximately 1000 nt from the HIV-1 5' region (termed E/DLS) including the encapsidation signal and the DIS-DLS . They found that the presence of an ectopic E/DLS near the 3' region of the genome resulted in the appearance of monomeric RNA in virus particles, suggesting that monomers can be packaged and that dimerization of HIV-1 genomic RNA is not required for packaging. However, they also found that two intact E/DLS regions had to be present on the same RNA molecule in order for packaging of monomers to occur. Therefore, it was assumed that these monomers had been generated from an intramolecular interaction between the two E/DLS regions. If we assume that such an intramolecular interaction between two DLS structures would occur on a single RNA molecule, however, might such a structure then not also appear as a dimer to a Gag protein that was attempting to package it? Although these data were interpreted to mean that dimerization is not required for packaging, they also suggest that some structure that is generated by the interaction of the two E/DLS regions might be recognized by Gag in order to facilitate packaging. In the context of wild-type genomic RNA containing only one E/DLS region, such a structure might then only be generated by an intermolecular interaction between two RNA molecules, i.e. a dimer. Hence, these results also imply that dimerization might be required for proper packaging.
|
15345057_p16
|
15345057
|
Can monomers be packaged?
| 4.431239 |
biomedical
|
Study
|
[
0.9992527365684509,
0.00045365834375843406,
0.0002935852389782667
] |
[
0.9988308548927307,
0.00034187856363132596,
0.0007155268685892224,
0.00011176627594977617
] |
en
| 0.999997 |
In a follow-up study, the same group created mutant HIV-1 particles that contained only monomeric RNAs, and concluded that these mutants demonstrated the complete separation of encapsidation from physical dimerization of retroviral RNA . However, they also reported that these viruses packaged only monomers, and that packaging efficiencies were approximately half those of wild-type, implying that dimerization is the sole mechanism to ensure the packaging of two copies of viral genomic RNA into each virus particle. In addition, the packaged monomers might have originally been weak dimers that dissociated during extraction and analysis, as has been pointed out in previous reports .
|
15345057_p17
|
15345057
|
Can monomers be packaged?
| 4.178677 |
biomedical
|
Study
|
[
0.9995850920677185,
0.00022637550137005746,
0.00018851962522603571
] |
[
0.9988234639167786,
0.00027891588979400694,
0.000824521470349282,
0.00007309346256079152
] |
en
| 0.999996 |
However, the above results do raise the issue of packaging specificity in mutant viruses. We and others have shown that, in COS cells, HIV-1 can incorporate significant amounts of spliced viral RNA when proper packaging of full-length viral genomic RNA is reduced . During assembly, Gag will always successfully package some RNA, and it is important to know the degree of specificity with which monomers versus dimers are packaged. If monomer-packaging mutants concomitantly package high levels of spliced viral RNA, then it is likely that packaging specificity may have been compromised by the existence of an extra E/DLS, and that the packaging of the monomers was non-specific. However, a lack of spliced viral RNA in these virions would indicate that the monomers were packaged with a high degree of specificity, and would have implications as to whether or not Gag initially recognizes viral genomic RNA in a dimeric versus a monomeric state. None of the viruses engineered to package only monomers were able to efficiently establish a new round of infection, suggesting that dimerization is required for replication if not for packaging. It is difficult to predict what other effects the addition of large segments of highly structured RNA might have on the viral life cycle.
|
15345057_p18
|
15345057
|
Can monomers be packaged?
| 4.35359 |
biomedical
|
Study
|
[
0.9993576407432556,
0.0003858240379486233,
0.0002564762835390866
] |
[
0.9986851811408997,
0.00041137560037896037,
0.0008019297383725643,
0.0001014601657516323
] |
en
| 0.999997 |
Another group reported similar phenotypes in the context of an HIV-1 mutant that was designed to have altered Gag/Gag-Pol ratios . Analysis of virion-derived genomic RNA from these viruses showed an increase in packaging of monomers, demonstrating that stable RNA dimers are not required for encapsidation of HIV-1 genomic RNA. Interestingly, these viruses also showed drastically reduced infectivity.
|
15345057_p19
|
15345057
|
Can monomers be packaged?
| 4.121794 |
biomedical
|
Study
|
[
0.9996139407157898,
0.0001944697869475931,
0.00019164006516803056
] |
[
0.9991557598114014,
0.00026060096570290625,
0.0005188012146390975,
0.00006476029375335202
] |
en
| 0.999994 |
We have also been studying the HIV-1 5' UTR and its putative interactions with Gag, and how these interactions affect dimerization and packaging activities. The DIS is known to be important for viral replication , reverse transcription , RNA dimerization , and packaging , as well as packaging specificity . However, despite the obvious importance of this stem-loop structure, work from our group has shown that defective viral replication caused by deletions in the DIS can be largely corrected by a series of compensatory point mutations identified in matrix, capsid, p2, and NC . These findings imply that the RNA sequences comprising the DIS interact in some way with these domains of Gag, and that when the RNA sequences are mutated, the virus will acquire adaptive mutations that potentially restore putative RNA-protein interactions over long-term culture. Since the originally deleted RNA sequences were in the DIS, we had naturally assumed that the major defect of these mutants would relate to RNA dimerization, and that compensatory mutations had arisen to correct defective RNA dimerization activity. To our surprise, this was not the case. Although our mutants did indeed yield reduced levels of dimerized genomic RNA in virus particles, the compensatory mutations in Gag that restored replication capacity did not correct dimerization defects . Rather, compensatory mutations apparently resulted in increased overall levels of viral genomic RNA that were packaged into virus particles, irrespective of impaired RNA dimerization. Similar effects on packaging were observed in the context of compensatory mutations identified during long-term culture of viruses containing mutations outside the DIS, such as the poly(A) hairpin and the U5-PBS complex , and between the PBS and SL1 . These findings again question the link between dimerization and packaging, since our compensatory point mutations were able to increase RNA packaging levels without correcting dimerization. One possibility is that the revertant viruses somehow gained the ability to package wild-type levels of RNA without correcting dimerization defects, i.e. they packaged more monomers. However, we also cannot rule out the possibility that our point mutations in Gag may have restored weak dimerization properties to the mutated RNAs, and that the latter dimers dissociated during extraction and analysis.
|
15345057_p20
|
15345057
|
Insights from forced evolution studies
| 4.481925 |
biomedical
|
Study
|
[
0.9990506768226624,
0.0006463098106905818,
0.0003029401123058051
] |
[
0.9989010095596313,
0.000366807944374159,
0.0005705869989469647,
0.00016158884682226926
] |
en
| 0.999996 |
In a follow-up study, we created two other DIS deletions and combined them with various combinations of the previously identified compensatory point mutations. We showed that these mutant viruses, ΔLoop (lacking the loop region of SL1) and ΔDIS (lacking the complete SL1) displayed defects in replication, RNA dimerization, and packaging. Once more, all of these but dimerization were largely corrected by the compensatory point mutations in Gag . Even a virus that lacked the DIS, e.g. ΔDIS, and which never showed any signs of viral growth in tissue culture, was able to replicate to significant extent when it also possessed the compensatory mutations.
|
15345057_p21
|
15345057
|
Insights from forced evolution studies
| 4.126334 |
biomedical
|
Study
|
[
0.9994505047798157,
0.00029483981779776514,
0.0002546414325479418
] |
[
0.9994418025016785,
0.0002088640903821215,
0.0002874851634260267,
0.00006181935168569908
] |
en
| 0.999997 |
The mechanism(s) whereby these compensatory point mutations functioned to restore replication had eluded us for some time. Recently, however, we employed an RNase protection assay to discriminate between genomic and spliced viral RNA packaged into virus particles. Our results showed that all of our 5' UTR mutant viruses aberrantly packaged increased levels of spliced viral RNA compared to wild-type virions. More importantly, however, the effect of one of our compensatory point mutations (i.e. MP2; a Thr->Ile substitution at position 12 of the SP1 spacer peptide in Gag) was to exclude spliced viral RNA from being packaged into mutant virions . Surprisingly, this single point mutation was also able to restore significant levels of virus replication to our ΔDIS mutant virus, which had been noninfectious in both T cell lines and blood mononuclear cells.
|
15345057_p22
|
15345057
|
Insights from forced evolution studies
| 4.229858 |
biomedical
|
Study
|
[
0.9995065927505493,
0.0003046928904950619,
0.00018876568356063217
] |
[
0.9993195533752441,
0.0002558411215431988,
0.00033343734685331583,
0.00009121331095229834
] |
en
| 0.999997 |
Previous work had suggested that the packaging of spliced viral RNA is a mechanism used by packaging mutants to fill the space that would normally be occupied by genomic RNA . Were this the case, then the MP2-mediated exclusion of spliced viral RNA from the virus particle should have been accompanied by increased packaging of genomic RNA. In the absence of MP2, the mutant particles contained lower levels of genomic RNA and higher levels of spliced viral RNA packaged than wild-type. In contrast, the presence of MP2 led to the exclusion of spliced viral RNA, but had no effect on packaging of genomic RNA. In the context of dimerization and packaging in the mutated viruses, it is possible that spliced viral RNAs, which do contain some RNA elements involved in RNA dimerization, including the DIS, might form heterodimers with molecules of genomic RNA. These putative heterodimers might be packageable, but it is unlikely that virions containing such genomes would be able to replicate, e.g. the noninfectious ΔDIS mutant. However, in the presence of MP2, the modified Gag protein might in some way block the formation of such an RNA heterodimer, thereby increasing the probability that dimers form between two genomic RNA molecules, resulting in partially restored levels of virus replication. Since these genomic RNA molecules are already mutated in dimerization signals, these weaker dimers would probably appear on a gel as monomers. In such a model, MP2 would act to restore dimerization, resulting in increased replication capacity, suggesting that dimerization is required for proper packaging to ensure that a particle is infectious. Unfortunately, this is virtually impossible to prove with current in vitro and in vivo protocols. New approaches to study dimerization and packaging within the cell will hopefully allow new hypotheses to be tested.
|
15345057_p23
|
15345057
|
Insights from forced evolution studies
| 4.531295 |
biomedical
|
Study
|
[
0.9992731213569641,
0.00047107721911743283,
0.0002557177795097232
] |
[
0.9956758618354797,
0.0007883008220233023,
0.003337210277095437,
0.00019859455642290413
] |
en
| 0.999996 |
The packaging of spliced viral RNA and/or the exclusion of such RNA species raises the question of whether the viral RNA sequence, or possibly the RNA structure, is important in proper assembly and/or structural integrity of the virus particle itself. Evidence in support of this possibility comes from studies on the binding of NC, in the context of full-length Gag, to viral genomic RNA. This might concentrate Gag proteins onto one or more RNA molecules, thereby facilitating Gag-Gag multimerization in a template-driven manner. Hence, viral genomic RNA would be a structural element, or scaffold, on which the virion can assemble . Other reports have shown that viral RNA can affect particle morphogenesis and structural stability , although the mechanisms involved are unclear. If RNA structure, or even the dimeric versus monomeric state of the RNA, truly does play a role in virion assembly and/or stability, this might also explain the apparent detection of monomeric RNA in the HIV-1 mutants mentioned above. For example, the duplication of large E/DLS sequences would undoubtedly have altered the overall structure of viral RNA, which might have resulted in the formation of unstable virus particles . Degradation of such particles could have indirectly caused the dissociation of dimers that would then appear as monomers on a gel. The fact that these viruses were all noninfectious may also have been due to the formation of unstable virus particles. Consistent with this concept, we found by electron microscopy that HIV-1 mutants lacking DIS stem sequences displayed an increased proportion of immature virus particles . This might mean that either the RNA structure, or the lack of a properly formed dimer, resulted in the production of virus particles with abnormal morphology. Since RNA can affect Gag cleavage, it is possible that mutations in the RNA might have also compromised the cleavage of Gag precursor proteins, which may subsequently have affected particle maturation . We believe that proper RNA dimerization may be a prerequisite for efficient virion assembly and structural stability.
|
15345057_p24
|
15345057
|
Insights from forced evolution studies
| 4.79698 |
biomedical
|
Study
|
[
0.998530387878418,
0.0010774371912702918,
0.0003920620074495673
] |
[
0.9959985017776489,
0.001198920770548284,
0.0022577825002372265,
0.0005448008305393159
] |
en
| 0.999996 |
As stated, the link between dimerization and packaging is a subject of ongoing debate , but we and others view dimerization as a prerequisite for packaging. Genomic RNA can be packaged as monomers , or alternatively as weak dimers that appear as monomers on gels, but mutant viruses that exhibit dimerization defects generally do not grow as well as wild-type viruses. The fact that our ΔDIS-MP2 virus can replicate in tissue culture, despite being severely compromised in genome dimerization, is evidence that efficient dimerization is not required for packaging or replication. In the absence of an authentic DIS, other sequences that affect dimerization may form a weak dimer that allows RNA to be recognized and adequately packaged . The contribution of the DIS might then be to significantly increase the efficiency of the dimerization process, resulting in more efficient packaging and replication. In conclusion, we agree with opinions expressed by others that the generation of virus particles able to package monomeric genomes is possible, but that dimerization is likely to be a prerequisite for the production of infectious viral progeny .
|
15345057_p25
|
15345057
|
Insights from forced evolution studies
| 4.392739 |
biomedical
|
Study
|
[
0.9993312358856201,
0.00042267495882697403,
0.00024617902818135917
] |
[
0.9970399737358093,
0.0005097283865325153,
0.002318727783858776,
0.00013144293916411698
] |
en
| 0.999997 |
It is clear that virus replication capacity is significantly affected whenever dimerization and/or packaging are compromised, suggesting that these activities can be exploited as anti-HIV drug targets. Indeed, the DIS was first proposed to be a potential therapeutic target at least 10 years ago, and antisense molecules were directed at this region of viral RNA , without practical outcome. Other approaches directly target the HIV-1 kissing-loop complex, which resembles the eubacterial 16S ribosomal aminoacyl-tRNA site, i.e. the target of aminoglycoside antibiotics such as paramycin and neomycin , both of which specifically bind to the kissing-loop complex. Drugs based on antibiotics with high affinity and specificity for the DIS may be a worthwhile approach, although efficacy might be compromised by the fact that HIV can replicate in the face of mutations that decrease genomic dimerization by more than 50% .
|
15345057_p26
|
15345057
|
The DIS as a therapeutic target?
| 4.296245 |
biomedical
|
Study
|
[
0.9992774128913879,
0.00040007781353779137,
0.0003225140681024641
] |
[
0.8110921382904053,
0.0016447767848148942,
0.18683397769927979,
0.0004291165969334543
] |
en
| 0.999998 |
RNA interference (RNAi) is a novel mechanism that regulates gene expression in which small interfering RNAs direct the targeted degradation of RNA in a sequence-specific manner (reviewed in Lee and Rossi ). Although RNAi is a powerful tool, it is not yet clear whether its therapeutic potential will materialize. This not-withstanding, several reports show that specific degradation of HIV-1 RNA is possible in infected cells , and reductions of p24 levels by as much as 4 logs have been achieved using RNAi directed against HIV-1 tat and rev . DNA vectors are currently being engineered that will allow for long-term production of siRNAs for use against chronic diseases, such as HIV-1.
|
15345057_p27
|
15345057
|
The DIS as a therapeutic target?
| 4.082061 |
biomedical
|
Review
|
[
0.998149037361145,
0.0009169672848656774,
0.0009340884280391037
] |
[
0.05782732367515564,
0.002347115660086274,
0.9393505454063416,
0.00047503761015832424
] |
en
| 0.999996 |
The DIS might also be a good candidate for sequence-specific targeting of HIV by RNAi therapy since it is highly conserved among naturally occurring virus isolates, and, due to its position upstream of the major splice donor, is contained in all HIV-1 RNA transcripts, both spliced and unspliced. Effective DIS-directed degradation of HIV RNA should confer the same viral phenotype as observed with our ΔDIS mutant, which never showed signs of virus replication in either permissive T cell lines or blood mononuclear cells . One concern with use of RNAi is how accessible certain RNA sequences might be. For example, complex secondary structures might cause some sequences to be buried and therefore inaccessible to the siRNA. However, this would not be a concern with DIS-directed RNAi, since the DIS contains a 6 nt palindromic sequence that is believed to initiate the dimerization process by binding to an identical sequence on another molecule of genomic RNA. If two 6 nt stretches of RNA can find each other on two 9200 nt strands of highly structured RNA, they should also be accessible to siRNAs.
|
15345057_p28
|
15345057
|
The DIS as a therapeutic target?
| 4.428926 |
biomedical
|
Study
|
[
0.9995452761650085,
0.0002683074271772057,
0.0001864097430370748
] |
[
0.997946560382843,
0.0009688178543001413,
0.0009547091322019696,
0.00012991443509235978
] |
en
| 0.999999 |
Recently, the practicality of RNAi-based therapies against HIV-1 was called into question when it was shown that HIV-1 was able to escape the antiviral pressure of RNAi by generating substitutions or even deletions within RNAi target sequences . This again highlights the versatility and plasticity of the HIV-1 genome. However, in these studies, the RNAi target sequences were located within the tat and nef genes, and the mutations that were generated blocked the effects of the RNAi without conferring any major detriment to virus replication. In contrast, RNAi may be more useful if targeted to more critical RNA elements within the genome, such as the DIS or the Ψ region, since any escape mutations that occur might result in viruses with severely impaired replication ability.
|
15345057_p29
|
15345057
|
The DIS as a therapeutic target?
| 4.255237 |
biomedical
|
Study
|
[
0.9995459914207458,
0.0002594215329736471,
0.00019463730859570205
] |
[
0.9802299737930298,
0.0006005843752063811,
0.019010504707694054,
0.0001589899038663134
] |
en
| 0.999997 |
All of these DIS-directed strategies rely on specifically targeting the viral RNA itself, which might not be practical given our inadequate knowledge of the overall structure of the HIV-1 5' region. The fact that RNA sequences such as SL1 and SL3 are known to form relevant RNA-protein interactions raises the possibility that the protein component of these interactions might also provide potential targets for anti-HIV therapy. Such approaches are currently being explored in research aimed at designing inhibitors of the TAR-Tat RNA-protein interaction . Similar approaches might also be developed to target RNA-protein interactions involving SL1 or SL3 and Gag.
|
15345057_p30
|
15345057
|
The DIS as a therapeutic target?
| 4.044554 |
biomedical
|
Study
|
[
0.9995704293251038,
0.00018006100435741246,
0.0002495285589247942
] |
[
0.942444920539856,
0.00479704886674881,
0.05248801037669182,
0.0002699527540244162
] |
en
| 0.999997 |
Current HIV combination therapies have demonstrated that a multi-targeted approach against the virus results in the greatest degree of suppression of virus replication. Therefore, the identification of novel targets for anti-HIV therapy could significantly improve HIV treatment strategies. HIV-1 RNA dimerization is clearly a critical event that could be exploited as a target once its complete mechanism is elucidated. It is pleasing to see that a number of laboratories that have actively researched RNA dimerization and packaging are now moving beyond conventional in vitro and in vivo approaches toward more biologically relevant methods. One group has taken chemical modification protocols commonly used for in vitro RNA analysis, and adapted them for use in virus-producing cells. Hence, structural analysis of viral RNA, that would previously be carried out only in vitro on short fragments of artificially transcribed RNA, can now be performed on in vivo -generated HIV-1 genomic RNA (J.-C. Paillart and R. Marquet, personal communication, and ). This method also allows comparisons of cellular and virion-derived HIV-1 RNA and represents a middle ground between classic in vitro and in vivo approaches. The goal of this work is to provide insight on the true structure of the HIV-1 leader, and on which RNA substructures are involved in dimerization. Preliminary data suggest that viral RNA may already be dimerized in the cytoplasm (J-C. Paillart and R. Marquet, unpublished data). This method might also have application in regard to in vivo foot-printing that could allow the study of RNA-protein interactions in the context of virus-producing cells.
|
15345057_p31
|
15345057
|
Future directions
| 4.288012 |
biomedical
|
Study
|
[
0.9995741248130798,
0.0002746738900896162,
0.0001512121089035645
] |
[
0.9973129630088806,
0.0005709370598196983,
0.0020024583209306,
0.00011361789074726403
] |
en
| 0.999995 |
The structure of the viral RNA that exists in the cell has long been a topic of interest, and recent data suggest that different RNA sequences might be involved in higher order intrastrand structures that favor the dimerization of the two RNA molecules. Such a model has been proposed , and is supported by numerous in vitro dimerization studies conducted on HIV-1, HIV-2, and SIV RNA . The model proposes that the HIV-1 5' UTR can form two alternating conformations, termed the long-distance interaction (LDI) and the branched multiple hairpin (BMH) structures. The LDI conformation is believed to exist when the RNA is in a monomer form, and is thought to form a long extended base-paired structure with almost all of the proposed stem-loop sequences buried. This structure is thought to be favored during certain steps of the life cycle, such as translation. In this model, NC has been shown to bind the LDI structure to induce a switch to the BMH structure , in which the DIS and ψ would then be exposed in a manner able to mediate dimerization and packaging. Such a 'riboswitch' is an attractive hypothesis, especially since similar mechanisms have recently been proposed to account for previously unexplained results in the field of gene regulation . Although there is currently little in vivo evidence directly supporting such a model in the case of retroviruses, the results of previous mutagenesis studies from several laboratories correlate with those that would be predicted from the riboswitch model, both concerning RNA packaging and RNA dimerization status . In regard to dimerization being a prerequisite for packaging, it would also be interesting to test whether an HIV-1 RNA molecule in the LDI conformation can be packaged. Since the BMH conformation is believed to mediate dimerization, one would assume that the LDI structures would not be packageable if dimerization is truly a packaging prerequisite.
|
15345057_p32
|
15345057
|
Future directions
| 4.785255 |
biomedical
|
Study
|
[
0.9987534284591675,
0.0007234435179270804,
0.0005231036921031773
] |
[
0.9882783889770508,
0.0010205189464613795,
0.010348523035645485,
0.000352535949787125
] |
en
| 0.999997 |
Others have developed a fluorescence resonance energy transfer (FRET)-based system to allow visualization of RNA-Gag interactions within cells (A.M. Lever and co-workers, unpublished data). Such a system might provide insight into the timing of genome selection and packaging. It will also be interesting to determine whether this system can be adapted to pinpoint how retroviral RNA dimerization takes place within cells, and whether dimerization indeed occurs before RNA is selected for packaging.
|
15345057_p33
|
15345057
|
Future directions
| 4.031249 |
biomedical
|
Study
|
[
0.9996919631958008,
0.0001249458728125319,
0.00018321890092920512
] |
[
0.9883819222450256,
0.00788543839007616,
0.0035274054389446974,
0.00020525010768324137
] |
en
| 0.999994 |
None declared.
|
15345057_p34
|
15345057
|
Competing interests
| 0.828596 |
other
|
Other
|
[
0.19056588411331177,
0.00526782963424921,
0.804166316986084
] |
[
0.017649337649345398,
0.97890704870224,
0.0020668196957558393,
0.001376785570755601
] |
it
| 0.999994 |
RSR gathered the information discussed in this review, and was primary author of the manuscript. CL and MAW carefully read the manuscript and offered insightful suggestions for its revision. All authors read and approved the final version.
|
15345057_p35
|
15345057
|
Author's contributions
| 0.885382 |
other
|
Other
|
[
0.02366505190730095,
0.003315198002383113,
0.9730197787284851
] |
[
0.0012306367279961705,
0.9903275370597839,
0.007131224498152733,
0.0013106607366353273
] |
en
| 0.999998 |
Prospective trials have the disadvantages of requiring a long time to complete, and using highly selected patient subgroups in tertiary centers. While one waits for the results to mature, this delays additional research to improve treatment. If there were a method that allowed earlier prediction of the results of prospective trials, advances in cancer treatment could be attained within a shorter time period.
|
15345027_p0
|
15345027
|
Background
| 3.377806 |
biomedical
|
Other
|
[
0.9952065348625183,
0.0028304557781666517,
0.0019629530142992735
] |
[
0.04908207803964615,
0.934022068977356,
0.01610792987048626,
0.0007879800396040082
] |
en
| 0.999997 |
There is a parametric lognormal model, proposed by Boag that had been retrospectively validated in the literature, and could be used prospectively for clinical trials to predict long-term survival rates several years earlier than would otherwise be possible using the standard life-table/actuarial Kaplan-Meier method of calculation .
|
15345027_p1
|
15345027
|
Background
| 3.661323 |
biomedical
|
Other
|
[
0.9975287318229675,
0.0005142769077792764,
0.0019569939468055964
] |
[
0.4254060983657837,
0.568265438079834,
0.005775462836027145,
0.0005530669004656374
] |
en
| 0.999997 |
The prognosis for metastatic breast cancer is generally poor and therefore it is believed that statistical prediction models for long-term survival rates are not needed. Nevertheless, specific subgroups of metastatic breast cancer patients exist, for which depending on the treatment given, the prognosis is improved so that some patients can survive for some time, particularly for those with limited organs involvement such as involvement with bone and/or skin only. In this situation, for which the present study was relevant, a prediction model, even for metastatic breast cancer, can be useful.
|
15345027_p2
|
15345027
|
Background
| 3.492557 |
biomedical
|
Study
|
[
0.998389482498169,
0.0006891363300383091,
0.0009213765151798725
] |
[
0.8164799213409424,
0.15615221858024597,
0.02645576372742653,
0.0009120451868511736
] |
en
| 0.999997 |
Breast cancer, among other cancers, has the highest incidence in women, and many studies are currently in progress to assess treatment regimens. If, even for a subgroup of patients, the 10- and 15-year survival rates can be predicted from follow-up data available only 3 years after a 5-year diagnosis period, this would be a useful means of obtaining study results earlier than would otherwise have been possible. For example, a 15-year survival rate calculated by the Kaplan-Meier method requires at least some patients to have been followed for 15 years. In addition prediction model such as the lognormal model can also be used to review the progress of treatment results for a specific period from a treatment center, and to compare that with another specific period of the same treatment center to evaluate the potential impact for any possible change in treatment policy or guideline.
|
15345027_p3
|
15345027
|
Background
| 3.962167 |
biomedical
|
Study
|
[
0.9994188547134399,
0.00022386461205314845,
0.0003572837158571929
] |
[
0.9712597131729126,
0.018566269427537918,
0.009983562864363194,
0.0001904515956994146
] |
en
| 0.999998 |
Boag's lognormal model for long-term cancer survival rates has been available for use for some 50 years. When the lognormal model was first proposed in the 1940s, it was difficult to implement because of a lack of computing power, and lack of good quality long-term follow-up data from cancer registries. Since 1970s the model has been used by some authors in breast cancer, cervix uteri cancer, head and neck cancer, intraocular melanoma, choroidal-ciliary body melanoma, and small cell lung cancer . Currently, although the computing power is sufficient, good quality follow-up data on a sufficient number of patients are seldom available, and it can be a limitation for its application. Large data registry such as the Surveillance, Epidemiology, and End Results (SEER) data with good long-term follow-up data available can overcome this potential limitation.
|
15345027_p4
|
15345027
|
Background
| 3.865157 |
biomedical
|
Review
|
[
0.9913076758384705,
0.0003769832255784422,
0.008315328508615494
] |
[
0.25640225410461426,
0.03913490101695061,
0.7040422558784485,
0.0004206524754408747
] |
en
| 0.999997 |
Between 1981 and 1985, 782 and 580 female patients of metastatic breast cancer were extracted respectively from the Connecticut and San Francisco-Oakland registries from the SEER database using SEER*Stat 5.0 software. The two registries were chosen because they are two of the earliest registries, with a large population. The data used were survival time, vital status, cause of death, age at diagnosis, and race.
|
15345027_p5
|
15345027
|
Methods
| 3.372553 |
biomedical
|
Study
|
[
0.9976192116737366,
0.0013302343431860209,
0.0010506163816899061
] |
[
0.9981599450111389,
0.001303258235566318,
0.0003785324515774846,
0.0001582731056259945
] |
en
| 0.999998 |
The cause-specific survival was defined as the interval from the date of diagnosis to the date of death from breast cancer or the last follow-up date for censoring purposes, if the patient was alive and still being followed at the time of analysis. The survival time of the uncured group of patients who died of breast cancer had been verified to follow a lognormal distribution previously .
|
15345027_p6
|
15345027
|
Methods
| 4.004687 |
biomedical
|
Study
|
[
0.9979022741317749,
0.0017670433735474944,
0.0003307453007437289
] |
[
0.9976826906204224,
0.0017800776986405253,
0.0003741919936146587,
0.0001631007471587509
] |
en
| 0.999997 |
Next, between 1991 and 1995, 752 and 632 female patients of metastatic breast cancer were extracted respectively from the two registries. The data were used to estimate the 15-year cause-specific survival rates before the year 2005. To be comparable, for both the 1981–1985 and 1991–1995 cohorts, the staging system used was the SEER historical system (classified as localized, regional, or distant, based on combined pathologic and clinical data). The choice of 1981–1985 and 1991–1995 has the advantage that the two time periods are not too far apart otherwise there would be too much changes of medical practice. These time periods have a minimum of 5 years follow-up.
|
15345027_p7
|
15345027
|
Methods
| 3.980576 |
biomedical
|
Study
|
[
0.9987144470214844,
0.0008152446243911982,
0.00047023387742228806
] |
[
0.9992823004722595,
0.0004004927759524435,
0.0002474294451531023,
0.00006979807221796364
] |
en
| 0.999998 |
The overall survival rates (OSR) of the two time periods were calculated using the Kaplan-Meier method. The actual relative survival rates (RSR) were calculated using SEER*Stat 5.0 software. The modified version of period analysis was applied using the Hakulinen method to obtain more up-to-date absolute survival rates (ASR) and relative survival rates (RSR) for comparison purpose with a computer program run by Microsoft Excel software.
|
15345027_p8
|
15345027
|
Methods
| 3.834886 |
biomedical
|
Study
|
[
0.9991801381111145,
0.00037730566691607237,
0.0004424524668138474
] |
[
0.9980498552322388,
0.0015531738754361868,
0.00031697694794274867,
0.00008001449168659747
] |
en
| 0.999996 |
The validation of the lognormal model has two phases. Phase 1 tests the goodness of fit to a lognormal distribution of the survival time of those cancer patients who died with their disease present, termed an uncured group with a fraction of 1-C, where C is the cured proportion of patients. The lognormal distribution is similar to the normal distribution in that if the variable in the normal is time t, the variable in the lognormal is the logarithm of t. In other words, the investigators attempt to show that the logarithm of the survival time follows a normal distribution. Phase 2 attempts to show that the lognormal model, using short-term follow-up data, can predict long-term survival rates comparable to those calculated by the Kaplan-Meier life-table method with long-term available. This model can be used to estimate long-term cause-specific survival rates (CSSR) by a maximum likelihood method (e.g., 10-year and 15-year survival rates) from only short-term follow-up data. The maximum likelihood method is used to estimate the CSSR at time τ, and is calculated as [C+(1-C)·Q]·100%, where Q is the integral of the lognormal distribution between the limits time τ and infinity.
|
15345027_p9
|
15345027
|
Validation of the lognormal model
| 4.153246 |
biomedical
|
Study
|
[
0.999452531337738,
0.00031610173755325377,
0.00023137283278629184
] |
[
0.9983469247817993,
0.0006918522994965315,
0.0008921692497096956,
0.00006915847188793123
] |
en
| 0.999997 |
The lognormal statistical model had been validated in stages III and IV breast cancer in a previous publication that survival rates could be estimated several years earlier than is possible using the standard life-table actuarial method . The survival time of unsuccessfully treated cases could be represented by a lognormal distribution, the long-term survival rates were predicted by Boag's method using a computer program run by Microsoft Excel. In this parametric lognormal model, the standard deviation S was fixed, and only the two remaining parameters, mean M and proportion cured C, were kept floating when using the maximum likelihood method. Multiple iterations converged to a stable solution for C.
|
15345027_p10
|
15345027
|
Validation of the lognormal model
| 4.096315 |
biomedical
|
Study
|
[
0.9995248317718506,
0.00021995762654114515,
0.00025519763585180044
] |
[
0.9991588592529297,
0.00035989118623547256,
0.0004267331678420305,
0.00005446181967272423
] |
en
| 0.999998 |
A 5-year period of diagnosis could be selected and patients followed as a cohort for an additional 3 years. The current study was for metastatic breast cancer patients treated between 1981 and 1985, with follow-up to the end of year 2000, making the series ideal for validating purposes. For example, for cases diagnosed during the 5-year period, prediction of the 15-year survival rate was made using data at the follow-up cutoff date of December 31, 1988 . The 15-year survival rate prediction was then validated by Kaplan-Meier life-table calculations using the follow-up data available in 2000.
|
15345027_p11
|
15345027
|
Validation of the lognormal model
| 4.037369 |
biomedical
|
Study
|
[
0.9991416931152344,
0.000557668216060847,
0.0003006011538673192
] |
[
0.9994115829467773,
0.00033894978696480393,
0.00018492322124075145,
0.00006452378147514537
] |
en
| 0.999999 |
For metastatic breast cancer patients treated between 1991 and 1995, and follow-up to the end of year 2000, prediction of the 15-year survival rate was made using data at the follow-up cutoff date of December 31, 1998 before the year 2005.
|
15345027_p12
|
15345027
|
Validation of the lognormal model
| 3.258805 |
biomedical
|
Study
|
[
0.9951189756393433,
0.003934892825782299,
0.0009461683221161366
] |
[
0.9944053888320923,
0.004840136505663395,
0.0004614319186657667,
0.0002930366899818182
] |
en
| 0.999995 |
From the cohort of 1981–1985 inclusively, 782 patients from the Connecticut registry were followed to the end of 1988. The lognormal model predicted the 15-year CSSR to be 7.1% (95% CI, 1.8–12.4). The 15-year CSSR was 8.3% (95% CI, 5.8–10.8) validated by the Kaplan-Meier calculation using actuarial follow-up data up to the end of year 2000.
|
15345027_p13
|
15345027
|
Results
| 4.102137 |
biomedical
|
Study
|
[
0.9983330368995667,
0.0013128428254276514,
0.0003541317128110677
] |
[
0.9990468621253967,
0.0005790079012513161,
0.00025947377434931695,
0.0001145561327575706
] |
en
| 0.999999 |
From the cohort of 1981–1985 inclusively, 580 patients from the San Francisco-Oakland registry were followed to the end of 1988. The lognormal model predicted the 15-year CSSR to be 9.2% (95% CI, 3.9–14.5). The 15-year CSSR was 7.0% (95% CI, 4.3–9.7) validated by the Kaplan-Meier calculation using actuarial follow-up data up to the end of year 2000.
|
15345027_p14
|
15345027
|
Results
| 4.094785 |
biomedical
|
Study
|
[
0.9984334111213684,
0.0011988029582425952,
0.00036783606628887355
] |
[
0.999110758304596,
0.0005388816352933645,
0.00024474188103340566,
0.00010563142859609798
] |
en
| 0.999998 |
Using the same method, the cohort of 1991–1995 inclusively, 752 patients from the Connecticut registry were followed to the end of 1998. The lognormal model predicted the 10-year CSSR to be 12.6% (95% CI, 7.3–17.9). The 10-year CSSR was 11.3% (95% CI, 7.8–14.8) validated by the Kaplan-Meier calculation using actuarial follow-up data up to the end of year 2000. The lognormal model predicted the 15-year CSSR to be 9.1% (95% CI, 3.8–14.4), which cannot be validated before 2005.
|
15345027_p15
|
15345027
|
Results
| 4.096001 |
biomedical
|
Study
|
[
0.9989468455314636,
0.0007106419070623815,
0.0003424746682867408
] |
[
0.9992424249649048,
0.0004456305759958923,
0.00023447454441338778,
0.00007756189734209329
] |
en
| 0.999997 |
For the cohort of 1991–1995 inclusively, 632 patients from the San Francisco-Oakland registry were followed to the end of 1998. The lognormal model predicted the 10-year CSSR to be 17.0% (95% CI, 12.1–21.9). The 10-year CSSR was 15.9% (95% CI, 11.4–20.4) validated by the Kaplan-Meier calculation using actuarial follow-up data up to the end of year 2000. The lognormal model predicted the 15-year CSSR to be 14.7% (95% CI, 9.8–19.6), which cannot be validated before 2005.
|
15345027_p16
|
15345027
|
Results
| 4.101727 |
biomedical
|
Study
|
[
0.9987836480140686,
0.0008535349043086171,
0.0003627518017310649
] |
[
0.9991845488548279,
0.0004959746147505939,
0.00023514640633948147,
0.00008429944136878476
] |
en
| 0.999998 |
For the period 1991–1995, there was not much change of only about 2% absolute percentage point in the predicted 15-year CSSR for the Connecticut registry, but there was an improvement of about 6% absolute percentage points for the San Francisco-Oakland registry when compared with the period 1981–1985 15-year CSSR, which was validated by the Kaplan-Meier calculation. (Table 1 )
|
15345027_p17
|
15345027
|
Results
| 3.704142 |
biomedical
|
Study
|
[
0.9975221753120422,
0.0009239311330020428,
0.001553861191496253
] |
[
0.9987988471984863,
0.0008331984281539917,
0.00029681585147045553,
0.00007116273627616465
] |
en
| 0.999996 |
For comparison purpose, the actual OSR and RSR were compared with the ASR and RSR obtained by the period analysis. (Tables 2 and 3 ) It was found that there were more patient survival improvements shown in the actual OSR and RSR for the San Francisco-Oakland registry, but not much for the Connecticut registry. However the period analysis results did not show such improvements.
|
15345027_p18
|
15345027
|
Results
| 2.286314 |
biomedical
|
Study
|
[
0.9863156676292419,
0.003732498735189438,
0.009951849468052387
] |
[
0.9946523904800415,
0.004531452897936106,
0.0005754934973083436,
0.0002405671402812004
] |
en
| 0.999997 |
Rutqvist studied the fit of Boag's lognormal model to the survival times of 8170 breast cancer cases reported to the Swedish Cancer Registry during 1961–1963. The model fitted the 1961–1963 data well for the entire case material and for patients aged less than 70 years. In this registry, the lognormal model did not fit the data for patients aged greater than 70 years, who were more likely to be censored because of coincidental causes of death. Another disadvantage stated by the author was that large number of patients was required to obtain estimates with reasonably small standard errors for breast cancer.
|
15345027_p19
|
15345027
|
Lognormal model
| 4.012377 |
biomedical
|
Study
|
[
0.9991961121559143,
0.00018650427227839828,
0.0006174097070470452
] |
[
0.9980670809745789,
0.0006395470118150115,
0.0012283952673897147,
0.00006501987809315324
] |
en
| 0.999997 |
With another series of the Norwegian Cancer Registry of 14,000 breast cancer cases, Rutqvist et al . deduced that lognormal is the best model because other models did not fit the observed survival in all stages, ages, and time periods (two-parameter models, such as exponential or extrapolated actuarial, or three-parameter models, such as sum of two exponential, exponential with shoulder, Weibull). Both the exponential and extrapolated actuarial models assume that the conditional relative survival is lowest immediately after treatment. With the lognormal model, the survival curve has a low initial mortality that rapidly increases to a maximum, with a slow decrease in the mortality after the maximum has occurred.
|
15345027_p20
|
15345027
|
Lognormal model
| 4.084731 |
biomedical
|
Study
|
[
0.9994019269943237,
0.0001710465585347265,
0.0004269581404514611
] |
[
0.9950273633003235,
0.0006655576289631426,
0.004238875117152929,
0.00006814548396505415
] |
en
| 0.999996 |
The lognormal model can only predict cause-specific survival, because other coincidental causes of death are too unpredictable (e.g., the rate of stroke). Therefore, overall survival cannot be predicted. The maximum likelihood method is the most accurate method for fitting the lognormal model with the smallest mean squared error. However, there are some requirements for its use. The maximum likelihood method fails to converge to a stable solution using the initial estimates if there is extensive censoring within the data. This occurs if patients are lost to follow-up or die from coincidental non-cancerous causes. The frequency of failure to yield a successful fit for lognormality was greater when one-fourth of cases were designated as lost to follow-up. Gamel et al . established a stable linear algorithm for fitting the lognormal model to survival data. To achieve convergence, some authors have fixed one or two parameters of the lognormal model to pre-selected values to simplify the iterative procedure required for convergence .
|
15345027_p21
|
15345027
|
Requirements for using the lognormal model
| 4.057822 |
biomedical
|
Study
|
[
0.9986456036567688,
0.0002466914302203804,
0.001107692369259894
] |
[
0.9850649237632751,
0.002041863277554512,
0.012794742360711098,
0.00009848835179582238
] |
en
| 0.999995 |
Prognostic factors in patients with distant metastases at the time of diagnosis were investigated by Rudan et al . , and Chapman et al . , primary tumor size was a significant prognostic factor. Engel et al . found that the number of metastatic cases and the time to metastasis depended on the tumor diameter at diagnosis. Cell growth is essential for the development of tumors. Tumor size is therefore the most important factor in describing tumor biology. As the tumor size increases, the probability of node-positivity increases. Another study group also found this correlation up to 5 cm . Tubiana and Koscielny have found a highly significant correlation between tumor size and the probability of distant metastasis. The distribution of tumor sizes at metastatic spread was lognormal with a median diameter equal to 3.5 cm. The patients were subdivided into 3 groups according to the histological grade. In each subgroup there was a significant correlation between tumor size and the probability of distant spread. The distributions were lognormal and the median size was markedly larger for grade 1 tumors.
|
15345027_p22
|
15345027
|
Some prognostic factors follow lognormal distribution
| 4.061481 |
biomedical
|
Study
|
[
0.999332845211029,
0.0003097860317211598,
0.0003573857247829437
] |
[
0.977135181427002,
0.00031085553928278387,
0.022415073588490486,
0.00013888553075958043
] |
en
| 0.999999 |
A number of quantitative postmortem observations regarding the size distribution of metastases have been published . These studies revealed a skewed distribution with a high proportion of smaller metastases, and a significant tail extending to the larger metastases, consistent with a lognormal distribution. The more detailed measurements from human liver metastases provided by Yamanami et al . were found to approximate the lognormal distribution reasonably well.
|
15345027_p23
|
15345027
|
Some prognostic factors follow lognormal distribution
| 4.049667 |
biomedical
|
Study
|
[
0.9996711015701294,
0.00011885931598953903,
0.00021011443459428847
] |
[
0.9932814836502075,
0.0005582971498370171,
0.006065791472792625,
0.00009436199616175145
] |
en
| 0.999997 |
A hypothesis was proposed by Kendal that the time available for the growth of metastases is normally distributed, presumably as a consequence of the summation of multiple independently distributed time intervals from each of the steps and of the Central Limit Theorem. For exponentially growing metastases, the corresponding size distribution would be lognormal; Gompertzian growth would imply a modified (Gompertz-normal) distribution, where larger metastases would occur less frequently as a consequence of a decreased growth rate. These two size distributions were evaluated against 18 human autopsy cases where precise size measurements had been collected from over 3900 macroscopic hematogenous organ metastases. The lognormal distribution provided an approximate agreement. Its main deficiency was a tendency to over-represent metastases greater than 10 mm diameter. These observations supported the hypothesis of normally distributed growth times, and qualified the utility of the lognormal and Gompertz-normal distributions for the size distribution of metastases.
|
15345027_p24
|
15345027
|
Some prognostic factors follow lognormal distribution
| 4.244864 |
biomedical
|
Study
|
[
0.9995576739311218,
0.0002384601248195395,
0.00020385101379361004
] |
[
0.9969111084938049,
0.0004440548364073038,
0.0025325019378215075,
0.0001123286274378188
] |
en
| 0.999997 |
Why is the lognormal model applicable to so many organ sites (Table 4 )? Boag's explanation for the lognormal survival time distribution was that if the patient was not cured by treatment, the length of the remaining survival time would be dependent principally on the growth rate of the tumor remnants. Pearlman estimated the growth rates of breast cancer that recurred in the scar, assuming that the recurrence started from a single cell. He found that the growth rates were approximately lognormally distributed. Likewise, von Fournier et al . found that the growth rates of breast cancers followed by serial mammography were lognormally distributed.
|
15345027_p25
|
15345027
|
Some prognostic factors follow lognormal distribution
| 4.055459 |
biomedical
|
Study
|
[
0.9993643164634705,
0.0001890311687020585,
0.0004466508107725531
] |
[
0.9583002924919128,
0.007845330983400345,
0.03361612558364868,
0.00023827023687772453
] |
en
| 0.999995 |
In order to determine whether current programs for the management of metastatic breast cancer have led to improved patient survival, Debonis et al . determined the median survival times for five-year intervals of 849 patients admitted to the City of Hope National Medical Center with metastatic breast cancer from 1955 to 1980. Survival times in each of the clinical subsets remained unchanged during the period of observation, regardless of the therapeutic modalities included in the treatment regimens. The study indicates that changes in palliative therapy for metastatic breast cancer during the 25 years of observation have not influenced overall survival. On the contrary, Dickman et al . studied the survival of cancer patients in Finland during the years 1955–1994. The 5-year RSR for distant metastases breast cancer had increased from 10% for the period 1955–1964 to 22% for 1985–1994.
|
15345027_p26
|
15345027
|
Variation of survival rates over time
| 4.040308 |
biomedical
|
Study
|
[
0.9992914199829102,
0.0004238566034473479,
0.0002846794668585062
] |
[
0.997080385684967,
0.00029651608201675117,
0.0025362737942487,
0.00008680936298333108
] |
en
| 0.999997 |
The tumor registry at Yale-New Haven Hospital, which began recording data in 1920, was utilized by Todd et al . to examine the ultimate outcome of all breast cancer patients who were initially diagnosed at Yale with metastatic breast cancer. The median survival of these patients increased steadily from 21 months in 1920 to 41 months in the decade from 1970 to 1980. The percentage of women actually surviving 5 years increased from 5% in the 1920s to approximately 25% in the 1960s. Despite the use of combination drug programs in the 1970s, the percentage of these patients remaining alive at 5 years remained near 25%. Firm conclusions cannot be made from a retrospective study spanning 60 years, although the trends depicted the lack of continued improvement indicate that the current therapeutic approach to metastatic breast cancer in that period may not result in dramatic improvement in overall survival.
|
15345027_p27
|
15345027
|
Variation of survival rates over time
| 4.083137 |
biomedical
|
Study
|
[
0.99886155128479,
0.0007875460432842374,
0.00035096038482151926
] |
[
0.9975988268852234,
0.0005508065805770457,
0.0017289536772295833,
0.00012147231609560549
] |
en
| 0.999998 |
Farrow et al . documented substantial geographical variation in patterns of treatment of cancer and other diseases. Because cancer treatment is not uniform nationwide in the States, survival following the diagnosis of cancer might also be expected to vary geographically. Survival data from the nine population-based registries in the SEER Program were analyzed for cancers of the stomach, colon, rectum, lung, breast, uterus, ovary, prostate, and bladder. The patients included all non-Hispanic white patients diagnosed with cancer of one of the selected sites during 1983–1991. Regional variation in crude five-year survival rates across the nine SEER areas was most marked for cancers of the uterus and prostate. For uterine cancer, for example, five-year survival ranged from 73.2% in Connecticut to 84.0% in Hawaii. Less marked variation was observed for cancers of the colon, rectum, and breast. For cancers of the bladder, ovary, stomach, and lung, survival rates five years after diagnosis were relatively invariant across the SEER areas.
|
15345027_p28
|
15345027
|
Geographical variation of survival rates
| 4.065958 |
biomedical
|
Study
|
[
0.9991846680641174,
0.00043843413004651666,
0.00037683016853407025
] |
[
0.9984458088874817,
0.000256966392043978,
0.0012257139896973968,
0.00007159180677263066
] |
en
| 0.999996 |
Maggard et al . also found that variations in the breast cancer mortality rates exist between states. A nearly 50% increase is observed between the states with the highest and lowest mortality rates. Adjusted analyses demonstrated that stage at presentation is a more important predictor of mortality variation than treatment differences. Goodwin et al . examined breast cancer incidence, survival, and mortality in the 66 health service areas covered by the SEER program for women aged 65 and older at diagnosis. They found that there was considerable geographic variation in survival from breast cancer among older women, and this contributed to variation in breast cancer mortality. The elevated mortality in the Northeast is apparent only in older women . For women aged 65 years and older, breast cancer mortality is 26% higher in New England than in the South, while incidence is only 3% higher. Breast cancer mortality for older women by state correlates poorly with incidence (r = 0.28).
|
15345027_p29
|
15345027
|
Geographical variation of survival rates
| 4.081024 |
biomedical
|
Study
|
[
0.9992561936378479,
0.0002836881030816585,
0.0004600742249749601
] |
[
0.9933820962905884,
0.00026107014855369925,
0.0062751080840826035,
0.00008174462709575891
] |
en
| 0.999996 |
Subsets and Splits
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