The present disclosure provides methods of analysing the nucleotide read sequences of a nucleic acid sample of interest using high throughput bidirectional sequencing. The methods of the present disclosure are designed to work even where bidirectional sequencing produces forward and reverse reads that are not of a sufficient read length to be paired via the complementary hybridisation of overlapping sequences at the 3° end of the sequence reads. The disclosure further provides computer-implemented methods, computer-readable storage mediums and devices that implement a method for preparing nucleic acid sequence results for analysis from non-overlapping sequence reads for screening a nucleic acid sample of interest for the expression of one or more target nucleotide sequences.

The present disclosure provides methods of analysing the nucleotide read sequences of a nucleic acid sample of interest using high throughput bidirectional sequencing. The methods of the present disclosure are designed to work even where bidirectional sequencing produces forward and reverse reads that are not of a sufficient read length to be paired via the complementary hybridisation of overlapping sequences at the 3° end of the sequence reads. The disclosure further provides computer-implemented methods, computer-readable storage mediums and devices that implement a method for preparing nucleic acid sequence results for analysis from non-overlapping sequence reads for screening a nucleic acid sample of interest for the expression of one or more target nucleotide sequences.

The present disclosure provides methods for predicting clinical response to therapy of a subject with an autoimmune disease or disorder and/or methods for predicting prognosis of a subject with immunodeficiency (e.g., leukemia) based on characterizing the B cell immune repertoire of the subject. In one aspect, methods provide for determining frequency of class switch recombination and/or somatic hypermutation of B cell receptor repertoires in samples prior to a treatment and predicting a subject's prognosis and/or response to treatment based on the measured class switching and/or somatic hypermutation frequency. In another aspect, methods provide for determination of immune receptor class switching and/or somatic hypermutation frequency and treating a subject based on the resulting prediction of prognosis and/or response to treatment based on the measured class switching and/or somatic hypermutation frequency.

The present disclosure provides methods for predicting clinical response to therapy of a subject with an autoimmune disease or disorder and/or methods for predicting prognosis of a subject with immunodeficiency (e.g., leukemia) based on characterizing the B cell immune repertoire of the subject. In one aspect, methods provide for determining frequency of class switch recombination and/or somatic hypermutation of B cell receptor repertoires in samples prior to a treatment and predicting a subject's prognosis and/or response to treatment based on the measured class switching and/or somatic hypermutation frequency. In another aspect, methods provide for determination of immune receptor class switching and/or somatic hypermutation frequency and treating a subject based on the resulting prediction of prognosis and/or response to treatment based on the measured class switching and/or somatic hypermutation frequency.

The present invention relates to a method for producing a medicament for the treatment or prevention of a tumor in a subject or a diagnostic agent for the detection of a tumor in a subject comprising (A) determining the expression of at least one nucleic acid molecule and/or at least one protein or peptide in a sample obtained from said subject, wherein the at least one nucleic acid molecule is selected from nucleic acid molecules (a) encoding a polypeptide comprising or consisting of the amino acid sequence of any one of SEQ ID NOs 1 to 5, (b) comprising or consisting of the nucleotide sequence of any one of SEQ ID NOs 6 to 10, (c) encoding a polypeptide which is at least 85% identical, preferably at least 90% identical, and most preferred at least 95% identical to the amino acid sequence of (a), (d) consisting of a nucleotide sequence which is at least 95% identical, preferably at least 96% identical, and most preferred at least 98% identical to the nucleotide sequence of (b), (e) consisting of a nucleotide sequence which is degenerate with respect to the nucleic acid molecule of (d), (f) consisting of a fragment of the nucleic acid molecule of any one of (a) to (e), said fragment comprising at least 150 nucleotides, preferably at least 300 nucleotides, more preferably at least 450 nucleotides, and most preferably at least 600 nucleotides, and (g) corresponding to the nucleic acid molecule of any one of (a) to (f), wherein T is replaced by U, and wherein the at least one protein or peptide is selected from proteins or peptides being encoded by the nucleic acid molecule of any one of (a) to (g); and (B) producing a medicament capable of inhibiting the expression of the at least nucleic acid molecule and/or the at least one protein or peptide in the subject, if the at least one nucleic acid molecule and/or at least one protein or peptide is expressed in (A), and/or (B′) producing a diagnostic agent capable of detecting in vivo the sites of expression of the at least nucleic acid molecule and/or the at least one protein or peptide in the subject, if the at least one nucleic acid molecule and/or at least one protein or peptide is expressed in (A).

The present invention relates to a method for producing a medicament for the treatment or prevention of a tumor in a subject or a diagnostic agent for the detection of a tumor in a subject comprising (A) determining the expression of at least one nucleic acid molecule and/or at least one protein or peptide in a sample obtained from said subject, wherein the at least one nucleic acid molecule is selected from nucleic acid molecules (a) encoding a polypeptide comprising or consisting of the amino acid sequence of any one of SEQ ID NOs 1 to 5, (b) comprising or consisting of the nucleotide sequence of any one of SEQ ID NOs 6 to 10, (c) encoding a polypeptide which is at least 85% identical, preferably at least 90% identical, and most preferred at least 95% identical to the amino acid sequence of (a), (d) consisting of a nucleotide sequence which is at least 95% identical, preferably at least 96% identical, and most preferred at least 98% identical to the nucleotide sequence of (b), (e) consisting of a nucleotide sequence which is degenerate with respect to the nucleic acid molecule of (d), (f) consisting of a fragment of the nucleic acid molecule of any one of (a) to (e), said fragment comprising at least 150 nucleotides, preferably at least 300 nucleotides, more preferably at least 450 nucleotides, and most preferably at least 600 nucleotides, and (g) corresponding to the nucleic acid molecule of any one of (a) to (f), wherein T is replaced by U, and wherein the at least one protein or peptide is selected from proteins or peptides being encoded by the nucleic acid molecule of any one of (a) to (g); and (B) producing a medicament capable of inhibiting the expression of the at least nucleic acid molecule and/or the at least one protein or peptide in the subject, if the at least one nucleic acid molecule and/or at least one protein or peptide is expressed in (A), and/or (B′) producing a diagnostic agent capable of detecting in vivo the sites of expression of the at least nucleic acid molecule and/or the at least one protein or peptide in the subject, if the at least one nucleic acid molecule and/or at least one protein or peptide is expressed in (A).

The present invention includes methods of detecting follicular regulatory T cells (T.sub.FR) comprising: obtaining a biological sample from a subject and detecting whether T.sub.FR are increased in the tumor sample by contacting the biological sample with antibodies that detect CD3.sup.+CD4.sup.+ FOXP3.sup.+BCL6.sup.+ T cells CD3.sup.+CD4.sup.+CXCR5.sup.+GITR.sup.+ T cells, or both, when compared to a healthy subject, and detecting the increase of T.sub.FR in the tumor sample. The present invention also includes combination therapy that depletes follicular regulatory T cells (T.sub.FR) with minimal effect on regulatory T cells (T.sub.REGS) to prevent or reduce immune related adverse effects (irAEs).

Provided in the present disclosure are a method for determining fetal nucleic acid concentration and a fetal genotyping method. According to the embodiments of the present disclosure, the method for determining cell-free fetal nucleic acid concentration includes: (1) acquiring sequencing data of a first nucleic acid sample of a pregnant woman and a reference genome sequence, the first nucleic acid sample of the pregnant woman containing cell-free fetal nucleic acids, and the sequencing data being composed of a plurality of sequencing reads; (2) selecting a predetermined region on the reference genome sequence and determining, based on the sequencing data of the first nucleic acid sample of the pregnant woman, mutation information in the predetermined region; and (3) determining the concentration of cell-free fetal nucleic acids corresponding to the predetermined region based on the mutation information in the predetermined region.

There is described herein a method of prognosing and/or predicting disease progression and/or in subject with prostate cancer, the method comprising: a) providing a sample containing mitochondrial genetic material from prostate cancer cells; b) sequencing the mitochondrial genetic material with respect to at least 1 patient biomarker selected from CSB1, OHR, ATP8 and HV1 (hypervariable region 1); c) comparing the sequence of the patient biomarkers to control or reference biomarkers to determine mitochondrial single nucleotide variations (mtSNVs); and d) determining the a prostate cancer prognosis; wherein a relatively worse outcome is associated with the presence of mtSNVs in CSB1, OHR, ATP8 and a relatively better outcome is associated with the presence of mtSNVs in HV1.

There is described herein a method of prognosing and/or predicting disease progression and/or in subject with prostate cancer, the method comprising: a) providing a sample containing mitochondrial genetic material from prostate cancer cells; b) sequencing the mitochondrial genetic material with respect to at least 1 patient biomarker selected from CSB1, OHR, ATP8 and HV1 (hypervariable region 1); c) comparing the sequence of the patient biomarkers to control or reference biomarkers to determine mitochondrial single nucleotide variations (mtSNVs); and d) determining the a prostate cancer prognosis; wherein a relatively worse outcome is associated with the presence of mtSNVs in CSB1, OHR, ATP8 and a relatively better outcome is associated with the presence of mtSNVs in HV1.