# Segment total copy numbers and allele B fractions using the Paired PSCBS method Segment total copy numbers and allele B fractions using the Paired PSCBS method [1]. This method requires matched normals. This is a low-level segmentation method. It is intended to be applied to one tumor-normal sample at the time. ```r ## Default S3 method: segmentByPairedPSCBS(CT, thetaT=NULL, thetaN=NULL, betaT=NULL, betaN=NULL, muN=NULL, rho=NULL, chromosome=0, x=NULL, alphaTCN=0.009, alphaDH=0.001, undoTCN=0, undoDH=0, ..., avgTCN=c("mean", "median"), avgDH=c("mean", "median"), flavor=c("tcn&dh", "tcn,dh", "sqrt(tcn),dh", "sqrt(tcn)&dh", "tcn"), tbn=is.null(rho), joinSegments=TRUE, knownSegments=NULL, dropMissingCT=TRUE, seed=NULL, verbose=FALSE, preserveScale=FALSE) ``` ## Arguments - `CT`: A `numeric` `vector` of J tumor total copy number (TCN) ratios in [0,+`Inf`) (due to noise, small negative values are also allowed). The TCN ratios are typically scaled such that copy-neutral diploid loci have a mean of two. - `thetaT, thetaN`: (alternative) As an alternative to specifying tumor TCN **ratios** relative to the match normal by argument `CT`, on may specify total tumor and normal signals seperately, in which case the TCN ratios `CT` are calculated as $CT = 2*thetaT/thetaN$. - `betaT`: A `numeric` `vector` of J tumor allele B fractions (BAFs) in [0,1] (due to noise, values may be slightly outside as well) or `NA` for non-polymorphic loci. - `betaN`: A `numeric` `vector` of J matched normal BAFs in [0,1] (due to noise, values may be slightly outside as well) or `NA` for non-polymorphic loci. - `muN`: An optional `numeric` `vector` of J genotype calls in {0,1/2,1} for AA, AB, and BB, respectively, and `NA` for non-polymorphic loci. If not given, they are estimated from the normal BAFs using `callNaiveGenotypes` as described in [2]. - `rho`: (alternative to `betaT` and `betaN`/`muN`) A `numeric` `vector` of J decrease-of-heterozygosity signals (DHs) in [0,1] (due to noise, values may be slightly larger than one as well). By definition, DH should be `NA` for homozygous loci and for non-polymorphic loci. - `chromosome`: (Optional) An `integer` scalar (or a `vector` of length J), which can be used to specify which chromosome each locus belongs to in case multiple chromosomes are segments. This argument is also used for annotation purposes. - `x`: Optional `numeric` `vector` of J genomic locations. If `NULL`, index locations `1:J` are used. - `alphaTCN, alphaDH`: The significance levels for segmenting total copy numbers (TCNs) and decrease-in-heterozygosity signals (DHs), respectively. - `undoTCN, undoDH`: Non-negative `numeric`s. If greater than 0, then a cleanup of segmentions post segmentation is done. See argument `undo` of `segmentByCBS`() for more details. - `avgTCN, avgDH`: A `character` string specifying how to calculating segment mean levels **after** change points have been identified. - `...`: Additional arguments passed to `segmentByCBS`(). - `flavor`: A `character` specifying what type of segmentation and calling algorithm to be used. - `tbn`: If `TRUE`, `betaT` is normalized before segmentation using the TumorBoost method [2], otherwise not. - `joinSegments`: If `TRUE`, there are no gaps between neighboring segments. If `FALSE`, the boundaries of a segment are defined by the support that the loci in the segments provides, i.e. there exist a locus at each end point of each segment. This also means that there is a gap between any neighboring segments, unless the change point is in the middle of multiple loci with the same position. The latter is what `DNAcopy::segment()` returns. - `knownSegments`: Optional `data.frame` specifying **non-overlapping** known segments. These segments must not share loci. See `findLargeGaps`() and `gapsToSegments`(). - `dropMissingCT`: If `TRUE`, loci for which 'CT' is missing are dropped, otherwise not. - `seed`: An (optional) `integer` specifying the random seed to be set before calling the segmentation method. The random seed is set to its original state when exiting. If `NULL`, it is not set. - `verbose`: See `Verbose`. - `preserveScale`: **Defunct - gives an error is specified.** ## Returns Returns the segmentation results as a `PairedPSCBS` object. ## Details Internally `segmentByCBS`() is used for segmentation. The Paired PSCBS segmentation method does **not** support weights. ## Reproducibility The "DNAcopy::segment" implementation of CBS uses approximation through random sampling for some estimates. Because of this, repeated calls using the same signals may result in slightly different results, unless the random seed is set/fixed. ## Whole-genome segmentation is preferred Although it is possible to segment each chromosome independently using Paired PSCBS, we strongly recommend to segment whole-genome (TCN,BAF) data at once. The reason for this is that downstream CN-state calling methods, such as the AB and the LOH callers, performs much better on whole-genome data. In fact, they may fail to provide valid calls if done chromosome by chromosome. ## Missing and non-finite values The total copy number signals as well as any optional positions must not contain missing values, i.e. `NA`s or `NaN`s. If there are any, an informative error is thrown. Allele B fractions may contain missing values, because such are interpreted as representing non-polymorphic loci. None of the input signals may have infinite values, i.e. -`Inf` or +`Inf`. If so, an informative error is thrown. ## Paired PSCBS with only genotypes If allele B fractions for the matched normal (`betaN`) are not available, but genotypes (`muN`) are, then it is possible to run a version of Paired PSCBS where TumorBoost normalization of the tumor allele B fractions is skipped. In order for this to work, argument `tbn` must be set to `FALSE`. ## Examples ```r verbose <- R.utils::Arguments$getVerbose(-10*interactive(), timestamp=TRUE) # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - # Load SNP microarray data # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - data <- PSCBS::exampleData("paired.chr01") str(data) # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - # Paired PSCBS segmentation # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - # Drop single-locus outliers dataS <- dropSegmentationOutliers(data) # Speed up example by segmenting fewer loci dataS <- dataS[seq(from=1, to=nrow(data), by=10),] str(dataS) R.oo::attachLocally(dataS) # Paired PSCBS segmentation fit <- segmentByPairedPSCBS(CT, betaT=betaT, betaN=betaN, chromosome=chromosome, x=x, seed=0xBEEF, verbose=verbose) print(fit) # Plot results plotTracks(fit) # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - # Bootstrap segment level estimates # (used by the AB caller, which, if skipped here, # will do it automatically) # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - fit <- bootstrapTCNandDHByRegion(fit, B=100, verbose=verbose) print(fit) plotTracks(fit) # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - # Calling segments in allelic balance (AB) # NOTE: Ideally, this should be done on whole-genome data # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - # Explicitly estimate the threshold in DH for calling AB # (which be done by default by the caller, if skipped here) deltaAB <- estimateDeltaAB(fit, flavor="qq(DH)", verbose=verbose) print(deltaAB) ## [1] 0.1657131 fit <- callAB(fit, delta=deltaAB, verbose=verbose) print(fit) plotTracks(fit) # Even if not explicitly specified, the estimated # threshold parameter is returned by the caller stopifnot(fit$params$deltaAB == deltaAB) # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - # Calling segments in loss-of-heterozygosity (LOH) # NOTE: Ideally, this should be done on whole-genome data # - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - # Explicitly estimate the threshold in C1 for calling LOH # (which be done by default by the caller, if skipped here) deltaLOH <- estimateDeltaLOH(fit, flavor="minC1|nonAB", verbose=verbose) print(deltaLOH) ## [1] 0.625175 fit <- callLOH(fit, delta=deltaLOH, verbose=verbose) print(fit) plotTracks(fit) # Even if not explicitly specified, the estimated # threshold parameter is returned by the caller stopifnot(fit$params$deltaLOH == deltaLOH) ``` ## Author(s) Henrik Bengtsson ## References [1] A.B. Olshen, H. Bengtsson, P. Neuvial, P.T. Spellman, R.A. Olshen, V.E. Seshan, **Parent-specific copy number in paired tumor-normal studies using circular binary segmentation**, Bioinformatics, 2011 [2] H. Bengtsson, P. Neuvial and T.P. Speed, **TumorBoost: Normalization of allele-specific tumor copy numbers from a single pair of tumor-normal genotyping microarrays**, BMC Bioinformatics, 2010 ## See Also Internally, `callNaiveGenotypes` is used to call naive genotypes, `normalizeTumorBoost` is used for TumorBoost normalization, and `segmentByCBS`() is used to segment TCN and DH separately. To segment tumor total copy numbers and allele B fractions **without** a matched normal, see `segmentByNonPairedPSCBS`(). To segment total copy-numbers, or any other unimodal signals, see `segmentByCBS`().

segmentByPairedPSCBS function