Oligonucleotide display processing device

Abstract

An oligonucleotide display processing device according to the present invention comprises: an input reception unit (34) that receives input of base information, sugar information, and phosphate group information of each of a plurality of nucleotides forming a target oligonucleotide, and arrangement information of the plurality of nucleotides; a storage unit (35) having pre-stored therein labels which each include at least one of a character or a symbol, and which are base labels corresponding to the base information, sugar labels corresponding to the sugar information, and phosphate group labels corresponding to the phosphate group information; an extraction unit (45) that extracts, from the storage unit, base labels, sugar labels, and phosphate group labels respectively corresponding to the base information, the sugar information, and the phosphate group information received by the input reception unit; and an output unit (46) that arranges, on the basis of the arrangement information of each of the nucleotides, the base labels, the sugar labels, and the phosphate group labels of the plurality of nucleotides extracted by the extraction unit, and disposes and displays, on a display unit (24), the base labels, the sugar labels, and the phosphate group labels of the respective nucleotides such that base labels of adjacent nucleotides are arranged side by side. The present invention enables easy understanding of the structure of oligonucleotides.

Description

Oligonucleotide Display Processing Device

[0001] The present invention relates to a display processing device for displaying the structure of an oligonucleotide.

[0002] Nucleic acid drugs consist of oligonucleotides, which are made up of several to several dozen nucleotides each having a base, sugar, and phosphate group, and can be chemically synthesized. Nucleic acid drugs are attracting attention as next-generation drugs because they can specifically act on intracellular molecules that cannot be targeted by conventional small molecule drugs or antibody drugs. Oligonucleotides used in nucleic acid drugs are often chemically modified on one or all of the base, sugar, and phosphate groups to improve in vivo stability and cell membrane permeability.

[0003] Confirming whether synthesized oligonucleotides have the intended sequence and modifications is important for ensuring their functionality as pharmaceuticals. Mass spectrometry is one method for confirming the sequence and presence or absence of modifications in oligonucleotides. In this method, mass spectrometry is first used to detect ion peaks related to the molecular weight of the oligonucleotide being analyzed, and molecular weight information is obtained. Next, the detected molecular weight-related ions are used as precursor ions, and dissociated using a predetermined method to generate multiple fragment ions, which are then mass analyzed to obtain a mass spectrum. By examining the assignment of each fragment peak observed in this mass spectrum, the partial structure of the oligonucleotide being analyzed can be determined, and the overall structure of the oligonucleotide can be estimated.

[0004] Thermo Fisher's "BioPharma Finder™" (Non-Patent Document 1) is known as computer software used for analyzing oligonucleotides. With this software, by inputting information about the structure (sequence and modifications) of the oligonucleotide to be confirmed via a keyboard or the like, letters or symbols corresponding to that information are displayed horizontally on the analysis screen. In addition, below the letters or symbols, colored bars indicating the structure corresponding to each fragment peak detected by the above-mentioned analytical method are displayed in association with the portion of the oligonucleotide. This makes it possible to see at a glance which portion of the oligonucleotide to be analyzed has had its structure determined.

[0005] The above character string is displayed using a common method for representing oligonucleotide structures. Specifically, the abbreviations representing the types of bases in each nucleotide of the oligonucleotide (adenine (A), thymine (T), guanine (G), cytosine (C), uracil (U), and hypoxanthine (H)) are arranged from the 5' end to the 3' end to represent the sequence. If the base, sugar, or phosphate group is modified, the abbreviations representing those modifications are added before, after, or before and after the abbreviation for the base. The abbreviations representing the modifications and the corresponding modification structures are stored as a dictionary by the software.

[0006] However, in this display method, the abbreviations indicating the sequence and the abbreviations indicating the modifications are displayed in the same column, making it difficult to extract the abbreviation indicating the sequence from the abbreviations at a glance, and making it difficult to understand the sequence of the oligonucleotide.

[0007] On the other hand, there is a method of representing the structure of an oligonucleotide by combining not only letters but also graphics or colors. For example, in Figure 1 of Non-Patent Document 2, letters indicating the type of base are each surrounded by a circle, connected by a line, and the structure of the oligonucleotide is represented by assigning colors to the letters, the interior of the circle, and the line to represent the base modification, sugar modification, and phosphate group modification, respectively. In this method, only the alphabet of the base is displayed as a letter, making it easy to recognize the base sequence of the oligonucleotide. However, since the modification of each part of the nucleotide constituting the oligonucleotide is represented only by color, it is difficult to understand which color corresponds to which modification.

[0008] "BioPharma Finder Software," [online], Thermo Fisher Scientific, [Retrieved November 14, 2022], Internet <URL: https: / / www.thermofisher.com / jp / ja / home / industrial / mass-spectrometry / liquid-chromatography-mass-spectrometry-lc-ms / lc-ms-software / multi-omics-data-analysis / biopharma-finder-software.html> Thomas C. Roberts, and 2 others, "Advances in oligonucleotide drug Delivery", Nature Reviews Drug Discovery, Springer Nature, 2020, 19, pp.673-694

[0009] The problem to be solved by the present invention is to provide an oligonucleotide display processing device that allows easy understanding of the structure (sequence and modification) of an oligonucleotide.

[0010] The oligonucleotide display processing device according to the present invention, which has been made to solve the above-mentioned problems, comprises: an input receiving unit that receives input of base information, sugar information, and phosphate group information for each of a plurality of nucleotides constituting a target oligonucleotide, as well as sequence information for the plurality of nucleotides; a memory unit in which base labels corresponding to the base information, sugar labels corresponding to the sugar information, and phosphate group labels corresponding to the phosphate group information, each of which is a label including at least one of a letter and a symbol, are pre-stored; an extraction unit that extracts from the memory unit the base labels, sugar labels, and phosphate group labels corresponding to the base information, sugar information, and phosphate group information, respectively, received by the input receiving unit; and an output unit that arranges the base labels, sugar labels, and phosphate group labels for each of the plurality of nucleotides extracted by the extraction unit based on the sequence information for each nucleotide, and displays the base labels, sugar labels, and phosphate group labels of each nucleotide on a display unit so that the base labels of adjacent nucleotides are aligned.

[0011] According to the oligonucleotide display processing device of the present invention, the base labels of each of the multiple nucleotides constituting an oligonucleotide are displayed in the order of the nucleotide sequence, with the base labels of adjacent nucleotides aligned side by side. In other words, since there are no sugar labels or phosphate group labels between the base labels of adjacent nucleotides, the base sequence of the oligonucleotide can be easily understood. Furthermore, since the sugar labels and phosphate group labels of each of the multiple nucleotides are displayed so as not to overlap with the rows of the base labels, the structure of the nucleotide, the presence or absence of modifications at each part of the nucleotide, etc. can be easily understood.

[0012] 1 is a block diagram showing a schematic configuration of a mass spectrometry system according to an embodiment of the present invention; FIG. 2 is a diagram showing an example of an oligonucleotide display screen in the mass spectrometry system;

[0013] Hereinafter, an embodiment of the present invention will be described with reference to the drawings.

[0014] 1 is a schematic diagram of a mass spectrometry system according to this embodiment. The mass spectrometry system includes a mass spectrometer 1 and a data processing and analysis device 2 that processes and analyzes mass analysis data acquired by the mass spectrometer 1.

[0015] The mass spectrometer 1 is preferably capable of performing mass analysis of fragment ions resulting from dissociation of ions derived from an object to be analyzed, such as in-source decay (ISD) analysis involving ISD and MS / MS analysis involving collision-induced dissociation (CID). For example, the mass spectrometer 1 may be a MALDI-TOFMS equipped with an ion source using matrix-assisted laser desorption / ionization (MALDI) and a time-of-flight mass separator (TOF) that separates various ions emitted from the ion source according to their mass-to-charge ratios, but is not limited thereto. The detection signal obtained by the mass spectrometer 1 is sent to a data processing / analysis device 2 via a communication line.

[0016] The data processing / analysis device 2 is actually a computer such as a workstation or personal computer, and is composed of a CPU (Central Processing Unit) 22, which is a central processing unit, a memory 23, a monitor (display unit) 24 consisting of an LCD (Liquid Crystal Display) or the like, an input operation unit 25 consisting of a keyboard, a mouse, etc., and a storage unit 30 consisting of a large-capacity storage device such as a hard disk or SSD (Solid State Drive), all of which are connected to one another.

[0017] The storage unit 30 stores an OS (Operating System) 37, a data processing and display program 32, and a compound library 33, and is also provided with an oligonucleotide structure storage unit 34, a label storage unit 35, and a measurement data storage unit 36. The compound library 33 stores structural formulas, mass spectra, and the like as information necessary for qualitative analysis of various types of compounds including oligonucleotides.

[0018] The data processing / analysis device 2 further includes an interface (I / F) 21 for direct connection with an external device or connection with an external device via a network such as a LAN (Local Area Network), and is connected to the mass spectrometer 1 via the I / F 21 via a USB cable. Note that, although the program for controlling the mass spectrometer 1 (MS control program) is assumed to be installed in the mass spectrometer 1, the MS control program may be installed on the same computer as the data processing / analysis device 2 or on a computer separate from that computer. The data processing / display program 32 may also be installed on a computer separate from that of the data processing / analysis device 2, in which case the computer is connected to the mass spectrometer 1 and the data processing / analysis device 2 via a USB cable.

[0019] 1 shows a mass spectrum creation unit 41, a peak extraction unit 42, a peak assignment determination unit 43, a peak assignment result analysis unit 44, a label extraction unit 45, and a display control unit 46, all of which are basically functional means realized in the form of software when the CPU 22 executes the data processing and display program 32.

[0020] In the mass spectrometry system of this embodiment, when mass spectrometry without fragmentation (MS1 analysis) and mass spectrometry with fragmentation (MS / MS analysis) are performed under predetermined conditions on a sample containing an oligonucleotide to be analyzed in the mass spectrometer 1, ions of a predetermined mass or mass range are detected sequentially, and the detection signals are sent to the data processing / analysis device 2. As a result, two-dimensional data consisting of mass-to-charge ratio (m / z) and signal intensity (ion intensity) is stored in the measurement data storage unit 36 ​​in the data processing / analysis device 2.

[0021] The data processing and display program 2 performs various processes on the data thus stored in the measurement data storage unit 36. For example, the mass spectrum creation unit 41 creates an MS1 ​​spectrum and an MS / MS spectrum representing the mass-to-charge ratio and intensity of ions based on the two-dimensional data. The mass spectra created in this manner are stored in the measurement data storage unit 36.

[0022] The peak extraction unit 42 extracts fragment peaks from the MS / MS spectrum created by the mass spectrum creation unit 41. The peak attribution determination unit 43 compares the MS1 spectrum created by the mass spectrum creation unit 41 with mass spectrum patterns stored in the compound library 33 to extract oligonucleotides with high similarity, and also obtains information on partial structures corresponding to the mass-to-charge ratios of each fragment peak extracted by the peak extraction unit 42 from the compound library 33. The information on partial structures includes information on one or more nucleotides constituting the partial structure (hereinafter referred to as "nucleotide information"), such as the types of bases, sugars, and phosphate groups constituting each nucleotide, and whether or not they are modified.

[0023] For example, if a partial structure corresponding to a peak includes three nucleotides arranged in the order of nucleotide 1, nucleotide 2, and nucleotide 3, the peak assignment determination unit 43 acquires, as partial structure information, nucleotide information arranged in the order of the nucleotide sequence, such as the types of base, sugar, and phosphate group of nucleotide 1 and whether or not they are modified, the types of base, sugar, and phosphate group of nucleotide 2 and whether or not they are modified, and the types of base, sugar, and phosphate group of nucleotide 3 and whether or not they are modified. The partial structure information acquired by the peak assignment determination unit 43 in this manner is stored in the oligonucleotide structure memory unit 34. The peak assignment result analysis unit 44 acquires the partial structure from the oligonucleotide structure memory unit 34 and analyzes the partial structure information according to a theoretical algorithm based on the known structure and mass of the nucleotides to estimate the structure of the target oligonucleotide. Information about the estimated oligonucleotide structure (i.e., information about the types and sequences of nucleotides constituting the oligonucleotide, and whether or not each nucleotide's base, sugar, and phosphate group is modified) is also stored in the oligonucleotide structure memory unit 34. In this embodiment, the oligonucleotide structure memory unit 34 corresponds to an input receiving unit.

[0024] Once the structure of the target oligonucleotide has been deduced in this manner, the label extraction unit 45 extracts labels corresponding to the bases, sugars, and phosphate groups of each of the multiple nucleotides constituting the oligonucleotide (referred to as base labels, sugar labels, and phosphate group labels, respectively), as well as labels corresponding to the types of modifications (hereinafter referred to as modification labels) if these bases, sugars, and phosphate groups are modified, from the label storage unit 35. The display control unit 46 creates display data (oligonucleotide display data) in which the various labels extracted by the label extraction unit 45 are laid out for each nucleotide based on the sequence (base sequence) of the nucleotides constituting the oligonucleotide, and displays an image showing the structure of the oligonucleotide on the monitor 24. In this embodiment, the display control unit 46 functions as an output unit.

[0025] In addition to such oligonucleotide display data, the display control unit 46 may obtain information on partial structures corresponding to each fragment peak from the oligonucleotide structure storage unit 34, and create display data for the partial structures to display on the monitor 24. For example, the partial structure display data may be data for displaying a bar of a length corresponding to the size of the partial structure in the oligonucleotide, in correspondence with the oligonucleotide. As a result, the monitor 24 displays the oligonucleotide structure and the assignment results of each fragment peak in correspondence with the oligonucleotide.

[0026] 2 shows an example of an analysis result display screen displayed on the monitor 24. In this example, the upper section of the analysis result display screen displays the molecular structure of the oligonucleotide being analyzed, the middle section displays the assignment results of the fragment peaks, and the lower section displays an explanation of the molecular structure and the assignment results. As can be seen from the display in the upper section of FIG. 2, in this embodiment, the molecular structure of the oligonucleotide is displayed as an image in which the labels (base label, sugar label, phosphate group label) corresponding to the bases, sugars, and phosphate groups contained in each of the multiple nucleotides constituting the oligonucleotide are grouped together for each nucleotide, and are arranged in the order of the nucleotides so that the base labels of adjacent nucleotides are aligned horizontally.

[0027] Here, the base label consists of a label with a single capital letter representing the base (A, G, C, T, U, H) surrounded by a hexagon, the sugar label consists of a pentagonal figure, and the phosphate group label consists of a label with a single lowercase letter representing the phosphate group (p, s) surrounded by a circle. If the base, sugar, or phosphate group is modified, each label contains a label indicating the modification. Among the labels representing the phosphate group, "p" indicates a phosphate group, and "s" indicates a thiophosphate. Furthermore, the label indicating the base modification consists of a single lowercase letter (e.g., m (methyl)). Furthermore, the label indicating the sugar modification consists of one or two lowercase letters and a symbol (e.g., d (2'-deoxy, DNA, no modification), r (2'-hydroxy, RNA, no modification), *(2'-O-methyl), e (2'-O-(methoxyethyl)), f (2'-fluoro, etc.). These base labels, sugar labels, and phosphate group labels are not limited to the examples described above and can be changed as appropriate. They may also be user-configurable.

[0028] In this embodiment, the sugar label is placed below the base label of each nucleotide, with a label indicating the sugar modification placed to the upper right of the sugar label and a phosphate group label placed to the lower left, but the sugar label, phosphate group label, and modification label may be placed in other positions. Furthermore, in this embodiment, the base labels and sugar labels are connected by solid lines, and adjacent nucleotides are connected by dashed lines, but either or both of these solid and dashed lines may be omitted.

[0029] Furthermore, the base labels, sugar labels, and phosphate group labels may be appropriately colored. For example, different colors may be assigned to the base labels for each type of base to facilitate understanding of the base sequence, or the label of a particular portion may be colored to highlight the structure of an impurity having a different structure (modification) from the main component.

[0030] Thus, an image representing the molecular structure of an oligonucleotide is composed of an image (hereinafter referred to as a labeled image) in which the base labels, sugar labels, and phosphate group labels are arranged and aligned according to the rules described above. Specifically, because there are no intervening labels between the base labels of adjacent nucleotides, it is easy to see that the base sequence of the oligonucleotide is 5'-CCTCTGGATTTGA-3'. Furthermore, because the sugar labels and phosphate group labels of each nucleotide are aligned so as not to overlap with the row of base labels, it is easy to see the structure of the nucleotide, the presence or absence of modifications at each part of the nucleotide, and so on, such as the replacement of phosphodiester bonds between all adjacent nucleotides with phosphorothioate bonds.

[0031] In addition, on the analysis result display screen shown in Figure 2, a bar indicating the assignment result of the fragment peak is displayed below the labeled image showing the molecular structure of the oligonucleotide. This bar has a length according to the size of the partial structure and is arranged corresponding to the position it occupies in the oligonucleotide. Therefore, it is possible to recognize which partial structure of the oligonucleotide has been confirmed from the length and position of the bar.

[0032] Aspects It will be apparent to those skilled in the art that the exemplary embodiments described above are examples of the following aspects.

[0033] (Item 1) An oligonucleotide display processing device according to one aspect of the present invention includes: an input receiving unit that receives input of base information, sugar information, and phosphate group information for each of a plurality of nucleotides constituting a target oligonucleotide, as well as sequence information for the plurality of nucleotides; a memory unit that pre-stores base labels corresponding to the base information, sugar labels corresponding to the sugar information, and phosphate group labels corresponding to the phosphate group information, each of which is a label including at least one of a letter and a symbol; an extraction unit that extracts from the memory the base labels, sugar labels, and phosphate group labels corresponding to the base information, sugar information, and phosphate group information, respectively, received by the input receiving unit; and an output unit that arranges the base labels, sugar labels, and phosphate group labels for each of the plurality of nucleotides extracted by the extraction unit based on the sequence information for each nucleotide, and displays the base labels, sugar labels, and phosphate group labels of each nucleotide on a display unit so that the base labels of adjacent nucleotides are aligned.

[0034] This allows the base labels of each of the multiple nucleotides constituting the oligonucleotide to be displayed in the order of the nucleotide sequence, with the base labels of adjacent nucleotides aligned side by side, i.e., without any intervening labels, making it easy to understand the sequence. Furthermore, the sugar label and the phosphate group label of each of the multiple nucleotides are displayed so as not to overlap with the line of the base labels, making it easy to understand the structure (modification) of each part of the nucleotide.

[0035] (2) In the oligonucleotide display processing device described in (1), the output unit may be configured so that the base labels of adjacent nucleotides are aligned horizontally, and the base label, sugar label, and phosphate group label of each nucleotide are arranged in order from top to bottom.

[0036] This makes it easy to understand the positional relationship between the base, sugar (the structure of the 2'-site of the sugar), and phosphate group attached to the pentose sugar of each nucleotide that makes up the oligonucleotide.

[0037] (Item 3) In the oligonucleotide display processing device according to item 1 or 2, at least one of the base label, the sugar label, and the phosphate group label may include a graphic.

[0038] This makes it easier to distinguish and recognize various signs.

[0039] (4) In the oligonucleotide display processing device described in paragraphs 1 to 3, at least one of the letters and symbols may be placed inside a predetermined figure, and the predetermined figure may be different from each other for the base label, the sugar label, and the phosphate group label.

[0040] This emphasizes the letters or symbols of the various signs, making them easier to distinguish and recognize.

[0041] REFERENCE SIGNS LIST 1 mass spectrometer 2 data processing / analysis device 22 CPU 24 monitor 23 memory 25 input operation unit 30 storage unit 32 data processing / display program 33 compound library 34 oligonucleotide structure storage unit 35 label storage unit 36 ​​measurement data storage unit 41 mass spectrum creation unit 42 peak extraction unit 43 peak assignment determination unit 44 peak assignment result analysis unit 45 label extraction unit 46 display control unit

Claims

**Claim 1**: A memory unit that pre-stores base labels, sugar labels, and phosphate group labels, which are labels containing at least one of characters and symbols corresponding to the bases, sugars, and phosphate groups of multiple types of nucleotides, an input reception unit that receives input of base information including information on the type of base and the presence or absence of modification in each of the multiple nucleotides constituting the oligonucleotide to be processed, sugar information including information on the type of sugar and the presence or absence of modification in the sugar, phosphate group information including information on the type of phosphate group and the presence or absence of modification in the phosphate group, and sequence information of the multiple nucleotides, an extraction unit that extracts, from the memory unit, base labels, sugar labels, and phosphate group labels corresponding to the base information, sugar information, and phosphate group information of the multiple nucleotides received by the input reception unit, an output unit that arranges the base labels, sugar labels, and phosphate group labels of each of the multiple nucleotides extracted by the extraction unit based on the sequence information for each nucleotide, and arranges and displays the base labels, sugar labels, and phosphate group labels of each nucleotide on a display unit so that the base labels of adjacent nucleotides are aligned, and when the output unit includes a nucleotide having a modification at any position of a base, a sugar, or a phosphate group in the multiple nucleotides received by the input reception unit, the output unit highlights and arranges on the display unit the label at the position having the modification among the base label, sugar label, and phosphate group label of the nucleotide. An oligonucleotide display processing device. **Claim 2** The oligonucleotide display processing device according to claim 1, wherein the output unit arranges the base labels of adjacent nucleotides side by side in the horizontal direction and arranges the base label, sugar label, and phosphate group label of each nucleotide in order from top to bottom. **Claim 3** The oligonucleotide display processing device according to claim 1, wherein at least one of the base label, sugar label, and phosphate group label includes a figure. **Claim 4** The oligonucleotide display processing device according to claim 1, wherein at least one of the character and the symbol is arranged inside a predetermined figure, and the predetermined figures are different from each other among the base label, sugar label, and phosphate group label. **Claim 5** A program that causes a computer to function as the oligonucleotide display processing apparatus according to any one of claims 1 to 4.