Method for predicting organ transplant rejection using next-generation sequencing

A method of predicting organ transplant rejection in a biological sample, obtained from a recipient who received an organ from a donor, by next-generation sequencing (NGS) or digital base amplification, the method comprising the steps of:

• obtaining a biological sample non-invasively, which contains donor-derived and recipient-derived cell-free nucleic acid molecules, from a recipient who received an organ from a donor;
• amplifying rs6490946, rs6490683, rs9552733, rs4770463, rs4770601, rs1886969, rs4771157, rs1927830, rs169332, rs524566, rs2764618, rs1832265, rs6562599, rs9544845, rs7236653, rs1284419, rs892430, rs17533, rs1786648, rs2419027, rs9949868, rs2090036, rs1893455, rs1593579, rs2222370, rs4798479, rs16951664, rs12606001, rs2821796, rs375484, rs2242661, rs4817371, rs2256417, rs413285, rs1014604, rs1102617, rs2183557, rs863075, rs2825608, rs12482714, rs2826390, rs2774494, rs7334546, rs9510597, rs9317882, rs943049, rs306395, rs1231048, rs9508327, rs9508716, rs798963, rs6561924, rs9564535, rs9572308, rs9574740, rs8095071, rs9303869, rs12327010, rs273696, rs8096542, rs2419041, rs8097023, rs8097433, rs9947829, rs4133291, rs2320747, rs10502290, rs602212, rs2027667, rs8134986, rs377191, rs2822661, rs1018676, rs2245411, rs2205533, rs2223079, rs8127266, rs1573442, rs1826318, rs2825610, rs2826117, rs2826392, rs9285110, rs6490713, rs9552874, rs9553022, rs9581121, rs9551233, rs9551406, rs2892463, rs7327256, rs2188584, rs9570290, rs1508494, rs7990298, rs7988209, rs1787013, rs2487257, rs716510, rs273701, rs4800786, rs2729429, rs11081004, rs1390431, rs8090831, rs9945284, rs4798412, rs2298583, rs8092926, rs329027, rs12482146, rs386838, rs11088040, rs926130, rs2736084, rs2823795, rs9978408, rs208932, rs2824762, rs7278137, rs128365, rs977660, rs2826396, rs876165, rs9506919, rs9805694, rs7992072, rs9553390, rs9512046, rs1778797, rs1570614, rs7986681, rs9534174, rs11839815, rs1876583, rs9542852, rs9575364, rs9952107, rs786018, rs6508351, rs9303900, rs16943649, rs17711702, rs878971, rs12967616, rs9967142, rs9967440, rs6506332, rs3786355, rs11660737, rs403093, rs431864, rs4588273, rs760345, rs2822965, rs975336, rs2823809, rs2226798, rs211962, rs8132870, rs2825560, rs2825688, rs1786427, rs2096905, rs4770032, rs9510334, rs2148443, rs736081, rs9551135, rs9507631, rs9508080, rs514669, rs914532, rs9534262, rs10507416, rs9564626, rs12586094, rs7323558, rs7231366, rs1030133, rs7228099, rs1785745, rs939139, rs9957591, rs1662304, rs12966492, rs1567612, rs6506138, rs694182, rs12185460, rs4239334, rs8087127, rs435081, rs2013669, rs467140, rs2822973, rs2823145, rs2823983, rs2824238, rs2824376, rs2211938, rs2825576, rs1474092, rs2826259, rs13433508, rs9509249, rs9510340, rs7323182, rs1830926, rs4770771, rs9512063, rs4573787, rs10478, rs7339250, rs4942486, rs12876644, rs9542105, rs9543171, rs7325529, rs7237577, rs968906, rs12961741, rs1785739, rs6508502, rs1026123, rs11081037, rs6507196, rs1145560, rs11872146, rs341237, rs16951141, rs8095514, rs1023868, rs439146, rs2822618, rs461853, rs2822975, rs2823152, rs2824133, rs13046156, rs243594, rs2825516, rs2825578, rs2825824, rs1786388, rs12483578, rs9509441, rs1834547, rs9510775, rs4770597, rs9507577, rs2497654, rs9579214, rs9506275, rs4943119, rs7330025, rs1361029, rs9592665, rs2031446, rs4796869, rs11080646, rs10210, rs1834545, rs8096605, rs11083386, rs336279, rs1790649, rs4799910, rs991045, rs4407150, rs9958938, rs7234111, rs7235989, rs2775537, rs416083, rs2822648, rs2822809, rs1002460, rs2823169, rs10470201, rs9984531, rs243601, rs1491658, rs2825583, rs377685, rs8128523, rs9786291, rs9786140, rs11096453, rs9786194, rs2058276, rs1358368, rs9786773, rs9786885, rs2571764, rs3096835, rs1864469, rs9786101, rs3991109, rs9785716, rs7067226, rs7067297, rs9785897, rs1276034, rs17307670, rs9786876, rs9785659, rs9786121, rs9786111, rs9786276, rs2161775, rs13304168, rs16980558, rs16980586, rs16980588, rs9786824, rs765557, rs9786386, rs9983568, rs8129332, rs229441, rs2826506, rs2014509, rs11096435, rs13304202 and rs2826399 marker sequences in cell-free nucleic acid molecules isolated from the biological sample;
• analyzing the amplified sequences by next-generation sequencing (NGS) or digital base amplification;
• based on the analysis of the sequences, determining the ratio between each of the donor-derived marker sequences and each of the recipient-derived marker sequences;
• and
• measuring the ratio over time, and predicting whether the recipient will have transplant rejection, graft dysfunction or organ failure when the ratio each of the donor-derived marker sequences increases.

A computer system comprising a computer readable medium encoded with a plurality of instructions for controlling a computing system to perform an operation of predicting organ transplant rejection in a biological sample, obtained from a recipient who received an organ from a donor, by use of next-generation sequencing (NGS) or digital base amplification,

• wherein the biological sample contains donor-derived and recipient-derived cell-free nucleic acid molecules from a recipient who received an organ from a donor, and
• wherein the operation comprises the steps of:
• receiving data obtained by analyzing rs6490946, rs6490683, rs9552733, rs4770463, rs4770601, rs1886969, rs4771157, rs1927830, rs169332, rs524566, rs2764618, rs1832265, rs6562599, rs9544845, rs7236653, rs1284419, rs892430, rs17533, rs1786648, rs2419027, rs9949868, rs2090036, rs1893455, rs1593579, rs2222370, rs4798479, rs16951664, rs12606001, rs2821796, rs375484, rs2242661, rs4817371, rs2256417, rs413285, rs1014604, rs1102617, rs2183557, rs863075, rs2825608, rs12482714, rs2826390, rs2774494, rs7334546, rs9510597, rs9317882, rs943049, rs306395, rs1231048, rs9508327, rs9508716, rs798963, rs6561924, rs9564535, rs9572308, rs9574740, rs8095071, rs9303869, rs12327010, rs273696, rs8096542, rs2419041, rs8097023, rs8097433, rs9947829, rs4133291, rs2320747, rs10502290, rs602212, rs2027667, rs8134986, rs377191, rs2822661, rs1018676, rs2245411, rs2205533, rs2223079, rs8127266, rs1573442, rs1826318, rs2825610, rs2826117, rs2826392, rs9285110, rs6490713, rs9552874, rs9553022, rs9581121, rs9551233, rs9551406, rs2892463, rs7327256, rs2188584, rs9570290, rs1508494, rs7990298, rs7988209, rs1787013, rs2487257, rs716510, rs273701, rs4800786, rs2729429, rs11081004, rs1390431, rs8090831, rs9945284, rs4798412, rs2298583, rs8092926, rs329027, rs12482146, rs386838, rs11088040, rs926130, rs2736084, rs2823795, rs9978408, rs208932, rs2824762, rs7278137, rs128365, rs977660, rs2826396, rs876165, rs9506919, rs9805694, rs7992072, rs9553390, rs9512046, rs1778797, rs1570614, rs7986681, rs9534174, rs11839815, rs1876583, rs9542852, rs9575364, rs9952107, rs786018, rs6508351, rs9303900, rs16943649, rs17711702, rs878971, rs12967616, rs9967142, rs9967440, rs6506332, rs3786355, rs11660737, rs403093, rs431864, rs4588273, rs760345, rs2822965, rs975336, rs2823809, rs2226798, rs211962, rs8132870, rs2825560, rs2825688, rs1786427, rs2096905, rs4770032, rs9510334, rs2148443, rs736081, rs9551135, rs9507631, rs9508080, rs514669, rs914532, rs9534262, rs10507416, rs9564626, rs12586094, rs7323558, rs7231366, rs1030133, rs7228099, rs1785745, rs939139, rs9957591, rs1662304, rs12966492, rs1567612, rs6506138, rs694182, rs12185460, rs4239334, rs8087127, rs435081, rs2013669, rs467140, rs2822973, rs2823145, rs2823983, rs2824238, rs2824376, rs2211938, rs2825576, rs1474092, rs2826259, rs13433508, rs9509249, rs9510340, rs7323182, rs1830926, rs4770771, rs9512063, rs4573787, rs10478, rs7339250, rs4942486, rs12876644, rs9542105, rs9543171, rs7325529, rs7237577, rs968906, rs12961741, rs1785739, rs6508502, rs1026123, rs11081037, rs6507196, rs1145560, rs11872146, rs341237, rs16951141, rs8095514, rs1023868, rs439146, rs2822618, rs461853, rs2822975, rs2823152, rs2824133, rs13046156, rs243594, rs2825516, rs2825578, rs2825824, rs1786388, rs12483578, rs9509441, rs1834547, rs9510775, rs4770597, rs9507577, rs2497654, rs9579214, rs9506275, rs4943119, rs7330025, rs1361029, rs9592665, rs2031446, rs4796869, rs11080646, rs10210, rs1834545, rs8096605, rs11083386, rs336279, rs1790649, rs4799910, rs991045, rs4407150, rs9958938, rs7234111, rs7235989, rs2775537, rs416083, rs2822648, rs2822809, rs1002460, rs2823169, rs10470201, rs9984531, rs243601, rs1491658, rs2825583, rs377685, rs8128523, rs9786291, rs9786140, rs11096453, rs9786194, rs2058276, rs1358368, rs9786773, rs9786885, rs2571764, rs3096835, rs1864469, rs9786101, rs3991109, rs9785716, rs7067226, rs7067297, rs9785897, rs1276034, rs17307670, rs9786876, rs9785659, rs9786121, rs9786111, rs9786276, rs2161775, rs13304168, rs16980558, rs16980586, rs16980588, rs9786824, rs765557, rs9786386, rs9983568, rs8129332, rs229441, rs2826506, rs2014509, rs11096435, rs13304202 and rs2826399 marker sequences, in the cell-free nucleic acid molecules isolated from the biological sample, by use of next-generation sequencing (NGS) or digital base amplification;
• based on the analysis of the sequences, determining the ratio between each of the donor-derived marker sequences and each of the recipient-derived marker sequences;
• comparing the ratio with one or more cutoff values;
• and
• based on the comparison, predicting whether or not organ transplant rejection in the recipient will be present.