Early detection of hepatocellular carcinoma in high risk populations using MALDI-TOF mass spectrometry
Hepatocellular carcinoma (HCC) is detected in a patient with liver disease. Mass spectrometry data from a blood-based sample from the patient is compared to a reference set of mass-spectrometry data from a multitude of other patients with liver disease, including patients with and without HCC, in a general purpose computer configured as a classifier. The classifier generates a class label, such as HCC or No HCC, for the test sample. A laboratory system for early detection of HCC in patients with liver disease is also disclosed. Alternative testing strategies using AFP measurement and a reference set for classification in the form of class-labeled mass spectral data from blood-based samples of lung cancer patients are also described, including multi-stage testing.
1. A method of generating a classifier for detecting HCC in a patient with liver disease, comprising the steps of:
a) conducting MALDI-TOF mass spectrometry on a set of blood-based samples from a multitude of patients with liver disease, including some patients with HCC and some patients without HCC;
b) storing a development set of mass spectrometry data as a result of conducting step a) on the set of blood-based samples, the development set of mass spectrometry data including feature values at a multitude of mass spectral features;
c) assigning a classification label to each member of the development set of step b) based on whether or not the patient associated with the sample had HCC;
d) separating the development set into training and test sets;
e) constructing a multitude of mini-classifiers using one or more of the feature values;
f) filtering the performance of the mini-classifiers operating on the training set and retaining only those mini-classifiers that meet a performance threshold;
g) generating a master classifier by combining the filtered mini-classifiers using a regularized combination method;
h) evaluating the performance of the master classifier on the test set;
i) repeating steps d), e), f), g) and h) for many different realizations of the separation of the development set into training and test sets; and
j) defining a final classifier from the master classifiers resulting from step g) and the repeated iterations of step i).
2. The method of claim 1 , wherein the development set of sample excludes those patients with high AFP expression levels.
3. The method of claim 1 , wherein the feature values include a feature value associated with AFP expression level measured in the blood-based samples of the development set.
4. The method of claim 1 , wherein the filtering step f) includes a sub-step of testing the performance of the mini-classifiers to classify spectra of healthy patients and retaining only those mini-classifiers whose healthy patient classification accuracy exceeds a pre-determined limit.
5. The method of claim 4 , wherein the method further comprises the step of varying the pre-determined limit and assessing master classifier performance with different values of the pre-determined limit.
6. The method of claim 1 , wherein the mini-classifiers implement a K-nearest neighbor algorithm and wherein the value of K is one of 5, 7, 9 and 11.
7. The method of claim 1 , the mini-classifiers perform classifications on the training set using single features, pairs of features, or single features, pairs of features and triplets of features in the set of feature values.
8. The method of claim 1 , wherein the mini-classifiers use at least 50 features in the set of features listed in Tables 1, 2, or 4:
TABLE 1
Left
Center
Right
3120.97
3132.28
3143.58
3144.12
3155.69
3167.27
3384.08
3395.16
3406.23
3408.04
3422.03
3436.02
3542.72
3558.63
3574.53
3582.98
3594.26
3605.53
3674.87
3686.35
3697.82
3760.14
3774.13
3788.12
3806.24
3818.82
3831.40
3856.97
3869.35
3881.73
3916.36
3928.94
3941.52
3943.13
3953.50
3963.87
4001.11
4015.00
4028.89
4040.17
4053.15
4066.14
4275.28
4290.71
4306.15
4875.22
4890.97
4906.72
4925.11
4937.26
4949.42
4978.09
4994.69
5011.30
5538.23
5560.98
5583.73
5617.26
5634.92
5652.58
5887.06
5905.52
5923.98
6052.67
6075.36
6098.05
6182.01
6205.21
6228.41
3033.39
3043.08
3052.77
3076.79
3088.26
3099.74
3100.53
3109.47
3118.41
3181.96
3188.28
3194.61
3196.98
3203.71
3210.45
3210.84
3219.94
3229.04
3229.67
3241.25
3252.83
3253.23
3263.29
3273.36
3273.56
3286.24
3298.93
3300.10
3312.52
3324.95
3325.11
3334.68
3344.26
3356.70
3369.99
3383.28
3436.83
3446.59
3456.35
3456.56
3465.51
3474.47
3498.50
3511.87
3525.24
3699.24
3708.03
3716.81
3788.32
3797.18
3806.04
3831.61
3843.99
3856.37
3882.14
3890.89
3899.65
3900.05
3907.70
3915.35
3964.20
3975.77
3987.33
4078.76
4093.64
4108.52
4114.45
4121.20
4127.94
4128.14
4135.99
4143.84
4204.62
4210.25
4215.88
4215.95
4221.58
4227.20
4230.26
4244.74
4259.22
4260.62
4267.38
4274.14
4306.57
4318.72
4330.88
4332.18
4341.37
4350.56
4351.18
4361.06
4370.94
4371.19
4378.76
4386.34
4386.42
4393.22
4400.02
4400.36
4409.29
4418.23
4418.48
4426.49
4434.51
4449.57
4456.23
4462.88
4462.92
4471.86
4480.79
4521.81
4529.99
4538.17
4538.34
4547.75
4557.17
4559.36
4570.55
4581.73
4581.81
4586.52
4591.23
4591.60
4600.18
4608.76
4618.52
4626.79
4635.05
4635.09
4646.56
4658.03
4668.49
4680.92
4693.35
4698.66
4711.22
4723.77
4748.42
4756.67
4764.91
4768.55
4775.84
4783.12
4783.33
4792.26
4801.20
4802.98
4818.65
4834.32
4846.65
4856.27
4865.90
4950.50
4962.87
4975.23
5011.71
5023.51
5035.31
5037.25
5045.01
5052.78
5052.88
5065.50
5078.12
5078.22
5086.70
5095.18
5095.28
5107.75
5120.21
5120.42
5129.61
5138.81
5139.22
5148.31
5157.40
5166.37
5176.18
5185.99
5186.62
5195.87
5205.12
5206.57
5222.14
5237.70
5260.04
5270.05
5280.07
5280.43
5289.47
5298.50
5317.72
5329.70
5341.67
5351.85
5360.73
5369.61
5396.95
5407.02
5417.10
5421.09
5430.67
5440.25
5440.61
5453.46
5466.31
5466.99
5474.16
5481.33
5482.34
5491.64
5500.94
5513.60
5522.04
5530.48
5662.56
5675.33
5688.10
5688.70
5706.56
5724.42
5724.99
5735.14
5745.28
5749.00
5762.00
5775.00
5769.52
5779.10
5788.68
5788.88
5797.16
5805.44
5814.82
5824.40
5833.98
5829.00
5841.50
5854.00
5846.35
5866.50
5886.66
5925.42
5936.84
5948.25
5978.65
5988.93
5999.21
6000.51
6008.02
6015.54
6015.97
6029.24
6042.51
6100.95
6109.32
6117.69
6117.83
6127.05
6136.28
6277.96
6285.65
6293.35
6293.42
6300.21
6306.99
6311.64
6325.69
6339.75
6392.91
6404.05
6415.19
6417.64
6433.26
6448.89
6449.43
6457.72
6466.01
6466.28
6476.88
6487.48
6488.92
6498.60
6508.28
6508.87
6529.73
6550.58
6579.32
6592.59
6605.86
6606.66
6626.41
6646.17
6646.57
6656.45
6666.32
6666.52
6676.70
6686.88
6687.20
6698.07
6708.94
6709.21
6728.64
6748.07
6785.46
6801.92
6818.39
6824.37
6835.85
6847.32
6847.72
6858.90
6870.07
6870.67
6881.05
6891.42
6891.62
6900.70
6909.78
6912.98
6920.66
6928.34
6926.00
6937.00
6948.00
6928.00
6942.50
6957.00
6950.89
6964.26
6977.63
6970.00
6973.50
6977.00
6977.77
6989.03
7000.29
7023.74
7040.65
7057.55
7058.93
7073.87
7088.82
7118.40
7141.12
7163.85
7174.99
7186.67
7198.34
7229.92
7240.81
7251.70
7252.50
7265.78
7279.07
7280.16
7293.73
7307.30
7345.41
7356.19
7366.97
7373.55
7387.02
7400.49
7401.58
7408.92
7416.25
7417.45
7426.93
7436.41
7436.81
7446.59
7456.37
7456.50
7478.98
7501.47
7506.46
7518.23
7530.00
7597.25
7614.12
7630.98
7694.84
7705.91
7716.99
7717.87
7734.18
7750.48
7751.84
7776.03
7800.21
7809.18
7826.04
7842.91
8126.48
8144.14
8161.80
8189.74
8205.00
8220.27
8249.00
8262.67
8276.34
8346.79
8372.73
8398.67
8400.30
8419.35
8438.40
8454.83
8472.27
8489.71
8491.46
8503.54
8515.61
8516.41
8526.59
8536.76
8536.96
8543.75
8550.53
8550.93
8574.58
8598.23
8608.21
8623.67
8639.14
8645.12
8655.40
8665.68
8666.08
8688.23
8710.38
8710.58
8731.23
8751.89
8752.28
8766.55
8780.82
8781.82
8803.67
8825.52
8826.12
8852.66
8879.20
8883.59
8893.87
8904.15
8904.35
8925.80
8947.25
8947.45
8958.92
8970.39
8984.28
8993.64
9003.00
9005.76
9023.07
9040.38
9042.84
9065.99
9089.14
9099.51
9129.35
9159.18
9159.58
9170.35
9181.13
9181.33
9190.11
9198.89
9199.09
9213.36
9227.63
9248.19
9256.96
9265.73
9266.58
9283.28
9299.98
9301.66
9319.42
9337.18
9337.38
9357.24
9377.09
9377.49
9389.47
9401.44
9401.64
9438.56
9475.48
9502.42
9523.27
9544.12
9553.30
9569.27
9585.23
9585.63
9596.71
9607.78
9608.18
9635.42
9662.66
9644.00
9655.00
9666.00
9688.60
9711.45
9734.30
9762.93
9794.85
9826.77
9828.25
9862.61
9896.96
9902.13
9925.28
9948.42
10190.81
10206.39
10221.97
10235.00
10256.74
10278.47
10316.61
10335.66
10354.72
10367.89
10390.04
10412.19
10424.76
10446.32
10467.87
10495.89
10507.13
10518.38
10518.60
10532.01
10545.41
10558.85
10574.44
10590.04
10604.17
10626.82
10649.46
10689.38
10720.71
10752.04
10757.69
10772.29
10786.89
10768.37
10777.35
10786.32
10809.51
10838.94
10868.38
10897.80
10916.69
10935.58
10983.52
11000.18
11016.85
11021.68
11043.26
11064.85
11089.22
11103.28
11117.33
11132.98
11147.56
11162.15
11277.36
11302.94
11328.52
11351.38
11368.14
11384.89
11415.69
11436.82
11457.95
11458.55
11476.97
11495.38
11501.17
11526.51
11551.86
11610.33
11627.89
11645.45
11656.23
11678.78
11701.33
11701.57
11726.48
11751.39
11757.73
11782.33
11806.93
11810.08
11827.25
11844.41
11856.78
11874.44
11892.10
11892.70
11908.27
11923.83
11924.03
11945.78
11967.53
12265.27
12293.61
12321.95
12421.12
12449.86
12478.60
12531.68
12560.61
12589.55
12590.15
12614.39
12638.64
12644.23
12668.08
12691.92
12711.88
12734.33
12756.78
12760.97
12780.53
12800.08
12806.50
12865.74
12924.98
12932.39
12965.13
12997.87
13015.80
13062.37
13108.95
13116.38
13129.95
13139.00
13143.92
13161.28
13178.64
13227.86
13240.86
13253.86
13290.59
13314.34
13338.09
13340.88
13360.63
13380.39
13387.24
13410.82
13434.40
13476.88
13509.03
13541.19
13542.36
13563.60
13584.84
13585.23
13605.30
13625.38
13686.96
13711.90
13736.85
13737.24
13758.09
13778.94
13779.33
13798.43
13817.53
13817.92
13836.43
13854.94
13855.33
13877.75
13900.16
13900.94
13926.08
13951.22
13927.00
13939.50
13952.00
13939.00
13954.00
13969.00
13952.77
13975.96
13999.16
13999.55
14031.90
14064.24
14065.80
14091.92
14118.03
14119.20
14146.87
14174.55
14176.89
14197.93
14218.98
14220.54
14248.21
14275.88
14395.63
14421.89
14448.15
14450.50
14479.92
14509.35
14510.52
14533.90
14557.29
14558.46
14585.94
14613.41
14752.16
14779.33
14806.50
14852.08
14877.14
14902.19
14939.25
14971.99
15004.73
16426.96
16511.73
16596.50
16598.45
16664.52
16730.58
16995.62
17026.80
17057.98
17104.30
17121.88
17139.47
17139.72
17155.18
17170.65
17176.14
17200.08
17224.03
17226.03
17267.43
17308.84
17341.77
17390.41
17439.05
17440.05
17472.35
17504.66
17568.51
17600.69
17632.87
17772.43
17807.18
17841.94
17852.95
17876.12
17899.29
17969.21
18022.01
18074.81
18226.45
18273.65
18320.85
18433.51
18489.23
18544.96
18549.22
18617.73
18686.25
18687.46
18726.44
18765.41
18766.02
18792.21
18818.40
18819.01
18856.46
18893.91
19049.56
19092.01
19134.45
19492.83
19552.08
19611.34
19882.92
19942.61
20002.29
20474.87
20549.17
20623.46
20711.16
20806.16
20901.17
20902.99
20955.06
21007.13
21008.35
21066.81
21125.28
21126.50
21174.91
21223.33
21224.54
21277.22
21329.90
21331.12
21383.19
21435.26
21651.45
21695.30
21739.15
21739.76
21763.51
21787.26
21787.70
21814.48
21841.25
TABLE 2
Feature definitions derived from
the 25k shot spectra
Left
Center
Right
3075.831
3085.942
3096.054
3100.514
3109.585
3118.655
3122.521
3129.956
3137.39
3189.73
3198.057
3206.384
3209.358
3217.685
3226.012
3234.338
3240.881
3247.423
3254.263
3261.252
3268.241
3276.865
3284.746
3292.626
3306.009
3315.079
3324.149
3387.195
3396.712
3406.228
3437.751
3446.227
3454.702
3544.513
3555.367
3566.222
3674.471
3684.731
3694.991
3808.295
3817.96
3827.625
3836.547
3844.279
3852.011
3883.236
3891.563
3899.89
3946.877
3953.717
3960.557
4003.678
4014.384
4025.09
4043.23
4051.557
4059.884
4240.695
4250.36
4260.025
4279.355
4290.507
4301.659
4334.372
4341.658
4348.944
4353.107
4361.732
4370.356
4373.032
4380.021
4387.009
4397.715
4407.232
4416.748
4419.722
4428.198
4436.673
4449.461
4463.884
4478.307
4562.17
4570.943
4579.716
4619.863
4626.554
4633.246
4636.22
4644.1
4651.981
4702.239
4711.756
4721.272
4767.962
4785.21
4802.459
4847.067
4854.947
4862.828
4882.456
4891.377
4900.299
4928.848
4936.729
4944.61
5056.724
5065.497
5074.27
5076.352
5084.084
5091.816
5095.385
5105.347
5115.31
5122.447
5130.03
5137.614
5139.695
5146.981
5154.267
5281.846
5290.619
5299.392
5398.719
5405.559
5412.399
5746.959
5763.167
5779.374
5790.08
5797.069
5804.057
5814.763
5822.347
5829.93
5834.688
5843.907
5853.126
5857.587
5866.062
5874.538
5880.188
5890.745
5901.303
6187.984
6196.608
6205.232
6277.794
6286.27
6294.745
6324.484
6332.067
6339.651
6407.752
6447.751
6487.749
6519.867
6530.87
6541.874
6601.054
6653.542
6706.031
6714.358
6727.443
6740.528
6794.95
6803.425
6811.901
6827.365
6837.476
6847.587
6850.859
6858.888
6866.917
6870.486
6881.489
6892.493
6912.12
6920.001
6927.882
6929.369
6941.562
6953.754
6955.836
6964.312
6972.787
7031.67
7044.606
7057.542
7177.984
7185.121
7192.259
7282.961
7293.519
7304.076
7378.72
7386.601
7394.482
7475.668
7485.184
7494.701
7551.799
7565.33
7578.861
7602.057
7615.44
7628.822
7661.237
7671.349
7681.46
7755.212
7765.025
7774.839
7813.202
7822.867
7832.532
7922.343
7939.145
7955.948
8007.098
8016.912
8026.726
8029.7
8042.636
8055.572
8134.082
8144.342
8154.602
8189.694
8204.117
8218.54
8402.92
8410.652
8418.384
8420.168
8428.049
8435.93
8517.711
8527.228
8536.744
8552.505
8561.576
8570.646
8573.917
8582.988
8592.058
8613.47
8624.473
8635.476
8647.074
8655.104
8663.133
8668.486
8686.627
8704.767
8709.823
8721.867
8733.911
8735.696
8743.428
8751.16
8754.728
8765.286
8775.843
8794.281
8809.745
8825.209
8835.023
8848.257
8861.49
8862.382
8870.858
8879.333
8901.935
8924.685
8947.435
8986.095
8994.571
9003.046
9007.21
9016.577
9025.945
9047.06
9066.985
9086.909
9109.213
9137.168
9165.122
9270.694
9282.144
9293.593
9306.083
9313.816
9321.548
9331.064
9353.071
9375.077
9400.355
9438.421
9476.486
9552.617
9568.527
9584.437
9613.284
9638.562
9663.84
9688.523
9716.328
9744.134
9901.749
9927.473
9953.197
10246.12
10256.98
10267.83
10330.28
10340.54
10350.8
10433.18
10445.67
10458.16
10516.45
10528.49
10540.54
10561.06
10575.78
10590.5
10820.38
10839.56
10858.74
11418.76
11437.81
11456.86
11494.96
11522.84
11550.72
11606.48
11626.69
11646.91
11660.85
11679.9
11698.95
11705.92
11733.8
11761.68
11764
11785.15
11806.29
11813.72
11825.8
11837.89
11861.12
11890.86
11920.6
11923.85
11944.76
11965.67
12279.32
12293.03
12306.73
12429.87
12450.08
12470.3
12546.97
12566.48
12586
12595.29
12607.61
12619.92
12657.09
12668.01
12678.93
12712.85
12727.72
12742.59
12830.41
12863.41
12896.4
12942.86
12962.38
12981.9
13051.13
13072.74
13094.34
13115.25
13127.57
13139.88
13145.46
13155.22
13164.97
13302.05
13315.99
13329.93
13598.04
13608.03
13618.02
13703.06
13719.09
13735.12
13742.55
13759.98
13777.4
13780.19
13792.04
13803.89
13827.12
13842.46
13857.79
13861.97
13881.72
13901.47
13910.3
13920.75
13931.21
13933.07
13942.59
13952.12
13958.16
13977.91
13997.66
14017.17
14036.22
14055.28
14074.79
14093.61
14112.43
14122.19
14144.49
14166.8
14174.69
14192.12
14209.54
14465.58
14482.54
14499.5
14515.3
14536.21
14557.12
14763.89
14779.46
14795.02
17010.56
17025.43
17040.29
17113.25
17140.43
17167.61
17235.45
17263.33
17291.21
17364.17
17387.87
17411.56
17446.41
17459.42
17472.43
17575.59
17597.89
17620.2
18258.65
18273.98
18289.32
18485.87
18500.28
18514.68
18561.15
18577.18
18593.21
18614.12
18634.8
18655.48
20824.08
20844.29
20864.5
20926.3
20957.2
20988.1
21035.5
21066.4
21097.3
21142.84
21168.39
21193.95
21242.28
21269.69
21297.11
22587.95
22611.65
22635.35
23003.36
23039.84
23076.32
TABLE 3
Feature Definitions of
Stable Features
Left
Center
Right
3350.649
3365.195
3379.741
3451.331
3462.145
3472.958
3473.429
3484.007
3494.586
3524.537
3550.157
3575.778
3662.644
3679.799
3696.955
3787.022
3810.493
3833.965
3940.773
3951.939
3963.106
3994.298
4013.968
4033.638
4034.299
4053.308
4072.317
4273.643
4289.346
4305.048
4327.528
4338.272
4349.016
4349.347
4359.264
4369.182
4394.717
4411.347
4427.977
4445.547
4462.738
4479.928
4614.584
4633.839
4653.095
4694.147
4712.329
4730.512
4762.248
4782.744
4803.24
4874.647
4890.184
4905.722
4923.904
4936.466
4949.028
5050.518
5064.072
5077.626
5090.188
5105.726
5121.263
5273.332
5287.713
5302.093
5394.988
5407.55
5420.112
5420.854
5430.492
5440.131
5685.241
5705.076
5724.912
5805.905
5821.442
5836.98
5889.874
5906.733
5923.593
5978.47
5988.109
5997.747
5998.452
6008.443
6018.434
6175.169
6192.194
6209.219
6263.104
6286.906
6310.708
6311.37
6332.692
6354.015
6403.978
6431.811
6459.644
6507.407
6528.233
6549.06
6596.536
6632.422
6668.308
6708.072
6729.89
6751.709
6821.129
6834.477
6847.825
6867.956
6879.772
6891.588
6909.094
6919.597
6930.1
6930.975
6946.292
6961.61
7173.396
7188.559
7203.722
7283.415
7297.402
7311.39
7369.242
7386.267
7403.292
7455.194
7472.219
7489.244
7542.786
7565.762
7588.738
7589.613
7612.37
7635.127
7801.647
7825.615
7849.582
7908.647
7934.905
7961.163
8180.855
8202.737
8224.619
8397.707
8408.638
8419.57
8419.805
8431.559
8443.313
8514.389
8527.447
8540.505
8748.112
8764.642
8781.171
9001.01
9015.225
9029.44
9097.259
9133.364
9169.468
9269.445
9285.148
9300.851
9548.459
9568.294
9588.129
9601.412
9638.392
9675.372
9678.435
9709.945
9741.455
9896.896
9931.938
9966.98
10316.41
10346.66
10376.91
10553.44
10577.74
10602.03
10602.7
10642.7
10682.7
10685.01
10719.72
10754.43
10803.36
10837.91
10872.45
11351.66
11371.84
11392.01
11504.11
11529.22
11554.33
11704.87
11727.35
11749.83
11917.43
11943.05
11968.67
12528.68
12564.22
12599.76
12813.98
12855.96
12897.95
12933.65
12960.43
12987.21
13745.24
13761.77
13778.3
13822.26
13839.45
13856.64
13858.63
13879.29
13899.95
13923.42
13937.64
13951.85
13955.16
13975.32
13995.49
14004.09
14035.33
14066.57
14069.21
14087.72
14106.24
18585.43
18631.04
18676.65
28026.88
28099.85
28172.82
28174.44
28213.97
28253.49
TABLE 4
Reduced Set of Features
Used in Classifiers
m/Z
3043
3109
3132
3286
3335
3708
3797
3891
3929
3954
4015
4053
4267
4291
4379
4530
4600
4627
4647
4757
4792
4891
5045
5065
5148
5196
5270
5474
5522
5561
5675
5779
5867
5906
6008
6286
6859
6881
6901
6937
6943
6964
6974
7041
7294
7409
7614
7826
8144
8504
8624
8731
8853
9066
9170
9190
9213
9257
9389
9439
9523
9655
9795
9863
10532
10627
10721
10839
10917
11437
11477
11726
11874
11946
13130
13161
13509
13564
13605
13712
13758
13798
13878
13940
13954
14032
14092
14147
14422
14480
14534
17122
17876
18274
18489
19552
20549
20955
21067
21175
“30000” (AFP).
9. The method of claim 1 , further comprising the step of repeating steps d) through j) for different numbers of features in the set of features listed in Table 1:
TABLE 1
Left
Center
Right
3120.97
3132.28
3143.58
3144.12
3155.69
3167.27
3384.08
3395.16
3406.23
3408.04
3422.03
3436.02
3542.72
3558.63
3574.53
3582.98
3594.26
3605.53
3674.87
3686.35
3697.82
3760.14
3774.13
3788.12
3806.24
3818.82
3831.40
3856.97
3869.35
3881.73
3916.36
3928.94
3941.52
3943.13
3953.50
3963.87
4001.11
4015.00
4028.89
4040.17
4053.15
4066.14
4275.28
4290.71
4306.15
4875.22
4890.97
4906.72
4925.11
4937.26
4949.42
4978.09
4994.69
5011.30
5538.23
5560.98
5583.73
5617.26
5634.92
5652.58
5887.06
5905.52
5923.98
6052.67
6075.36
6098.05
6182.01
6205.21
6228.41
3033.39
3043.08
3052.77
3076.79
3088.26
3099.74
3100.53
3109.47
3118.41
3181.96
3188.28
3194.61
3196.98
3203.71
3210.45
3210.84
3219.94
3229.04
3229.67
3241.25
3252.83
3253.23
3263.29
3273.36
3273.56
3286.24
3298.93
3300.10
3312.52
3324.95
3325.11
3334.68
3344.26
3356.70
3369.99
3383.28
3436.83
3446.59
3456.35
3456.56
3465.51
3474.47
3498.50
3511.87
3525.24
3699.24
3708.03
3716.81
3788.32
3797.18
3806.04
3831.61
3843.99
3856.37
3882.14
3890.89
3899.65
3900.05
3907.70
3915.35
3964.20
3975.77
3987.33
4078.76
4093.64
4108.52
4114.45
4121.20
4127.94
4128.14
4135.99
4143.84
4204.62
4210.25
4215.88
4215.95
4221.58
4227.20
4230.26
4244.74
4259.22
4260.62
4267.38
4274.14
4306.57
4318.72
4330.88
4332.18
4341.37
4350.56
4351.18
4361.06
4370.94
4371.19
4378.76
4386.34
4386.42
4393.22
4400.02
4400.36
4409.29
4418.23
4418.48
4426.49
4434.51
4449.57
4456.23
4462.88
4462.92
4471.86
4480.79
4521.81
4529.99
4538.17
4538.34
4547.75
4557.17
4559.36
4570.55
4581.73
4581.81
4586.52
4591.23
4591.60
4600.18
4608.76
4618.52
4626.79
4635.05
4635.09
4646.56
4658.03
4668.49
4680.92
4693.35
4698.66
4711.22
4723.77
4748.42
4756.67
4764.91
4768.55
4775.84
4783.12
4783.33
4792.26
4801.20
4802.98
4818.65
4834.32
4846.65
4856.27
4865.90
4950.50
4962.87
4975.23
5011.71
5023.51
5035.31
5037.25
5045.01
5052.78
5052.88
5065.50
5078.12
5078.22
5086.70
5095.18
5095.28
5107.75
5120.21
5120.42
5129.61
5138.81
5139.22
5148.31
5157.40
5166.37
5176.18
5185.99
5186.62
5195.87
5205.12
5206.57
5222.14
5237.70
5260.04
5270.05
5280.07
5280.43
5289.47
5298.50
5317.72
5329.70
5341.67
5351.85
5360.73
5369.61
5396.95
5407.02
5417.10
5421.09
5430.67
5440.25
5440.61
5453.46
5466.31
5466.99
5474.16
5481.33
5482.34
5491.64
5500.94
5513.60
5522.04
5530.48
5662.56
5675.33
5688.10
5688.70
5706.56
5724.42
5724.99
5735.14
5745.28
5749.00
5762.00
5775.00
5769.52
5779.10
5788.68
5788.88
5797.16
5805.44
5814.82
5824.40
5833.98
5829.00
5841.50
5854.00
5846.35
5866.50
5886.66
5925.42
5936.84
5948.25
5978.65
5988.93
5999.21
6000.51
6008.02
6015.54
6015.97
6029.24
6042.51
6100.95
6109.32
6117.69
6117.83
6127.05
6136.28
6277.96
6285.65
6293.35
6293.42
6300.21
6306.99
6311.64
6325.69
6339.75
6392.91
6404.05
6415.19
6417.64
6433.26
6448.89
6449.43
6457.72
6466.01
6466.28
6476.88
6487.48
6488.92
6498.60
6508.28
6508.87
6529.73
6550.58
6579.32
6592.59
6605.86
6606.66
6626.41
6646.17
6646.57
6656.45
6666.32
6666.52
6676.70
6686.88
6687.20
6698.07
6708.94
6709.21
6728.64
6748.07
6785.46
6801.92
6818.39
6824.37
6835.85
6847.32
6847.72
6858.90
6870.07
6870.67
6881.05
6891.42
6891.62
6900.70
6909.78
6912.98
6920.66
6928.34
6926.00
6937.00
6948.00
6928.00
6942.50
6957.00
6950.89
6964.26
6977.63
6970.00
6973.50
6977.00
6977.77
6989.03
7000.29
7023.74
7040.65
7057.55
7058.93
7073.87
7088.82
7118.40
7141.12
7163.85
7174.99
7186.67
7198.34
7229.92
7240.81
7251.70
7252.50
7265.78
7279.07
7280.16
7293.73
7307.30
7345.41
7356.19
7366.97
7373.55
7387.02
7400.49
7401.58
7408.92
7416.25
7417.45
7426.93
7436.41
7436.81
7446.59
7456.37
7456.50
7478.98
7501.47
7506.46
7518.23
7530.00
7597.25
7614.12
7630.98
7694.84
7705.91
7716.99
7717.87
7734.18
7750.48
7751.84
7776.03
7800.21
7809.18
7826.04
7842.91
8126.48
8144.14
8161.80
8189.74
8205.00
8220.27
8249.00
8262.67
8276.34
8346.79
8372.73
8398.67
8400.30
8419.35
8438.40
8454.83
8472.27
8489.71
8491.46
8503.54
8515.61
8516.41
8526.59
8536.76
8536.96
8543.75
8550.53
8550.93
8574.58
8598.23
8608.21
8623.67
8639.14
8645.12
8655.40
8665.68
8666.08
8688.23
8710.38
8710.58
8731.23
8751.89
8752.28
8766.55
8780.82
8781.82
8803.67
8825.52
8826.12
8852.66
8879.20
8883.59
8893.87
8904.15
8904.35
8925.80
8947.25
8947.45
8958.92
8970.39
8984.28
8993.64
9003.00
9005.76
9023.07
9040.38
9042.84
9065.99
9089.14
9099.51
9129.35
9159.18
9159.58
9170.35
9181.13
9181.33
9190.11
9198.89
9199.09
9213.36
9227.63
9248.19
9256.96
9265.73
9266.58
9283.28
9299.98
9301.66
9319.42
9337.18
9337.38
9357.24
9377.09
9377.49
9389.47
9401.44
9401.64
9438.56
9475.48
9502.42
9523.27
9544.12
9553.30
9569.27
9585.23
9585.63
9596.71
9607.78
9608.18
9635.42
9662.66
9644.00
9655.00
9666.00
9688.60
9711.45
9734.30
9762.93
9794.85
9826.77
9828.25
9862.61
9896.96
9902.13
9925.28
9948.42
10190.81
10206.39
10221.97
10235.00
10256.74
10278.47
10316.61
10335.66
10354.72
10367.89
10390.04
10412.19
10424.76
10446.32
10467.87
10495.89
10507.13
10518.38
10518.60
10532.01
10545.41
10558.85
10574.44
10590.04
10604.17
10626.82
10649.46
10689.38
10720.71
10752.04
10757.69
10772.29
10786.89
10768.37
10777.35
10786.32
10809.51
10838.94
10868.38
10897.80
10916.69
10935.58
10983.52
11000.18
11016.85
11021.68
11043.26
11064.85
11089.22
11103.28
11117.33
11132.98
11147.56
11162.15
11277.36
11302.94
11328.52
11351.38
11368.14
11384.89
11415.69
11436.82
11457.95
11458.55
11476.97
11495.38
11501.17
11526.51
11551.86
11610.33
11627.89
11645.45
11656.23
11678.78
11701.33
11701.57
11726.48
11751.39
11757.73
11782.33
11806.93
11810.08
11827.25
11844.41
11856.78
11874.44
11892.10
11892.70
11908.27
11923.83
11924.03
11945.78
11967.53
12265.27
12293.61
12321.95
12421.12
12449.86
12478.60
12531.68
12560.61
12589.55
12590.15
12614.39
12638.64
12644.23
12668.08
12691.92
12711.88
12734.33
12756.78
12760.97
12780.53
12800.08
12806.50
12865.74
12924.98
12932.39
12965.13
12997.87
13015.80
13062.37
13108.95
13116.38
13129.95
13139.00
13143.92
13161.28
13178.64
13227.86
13240.86
13253.86
13290.59
13314.34
13338.09
13340.88
13360.63
13380.39
13387.24
13410.82
13434.40
13476.88
13509.03
13541.19
13542.36
13563.60
13584.84
13585.23
13605.30
13625.38
13686.96
13711.90
13736.85
13737.24
13758.09
13778.94
13779.33
13798.43
13817.53
13817.92
13836.43
13854.94
13855.33
13877.75
13900.16
13900.94
13926.08
13951.22
13927.00
13939.50
13952.00
13939.00
13954.00
13969.00
13952.77
13975.96
13999.16
13999.55
14031.90
14064.24
14065.80
14091.92
14118.03
14119.20
14146.87
14174.55
14176.89
14197.93
14218.98
14220.54
14248.21
14275.88
14395.63
14421.89
14448.15
14450.50
14479.92
14509.35
14510.52
14533.90
14557.29
14558.46
14585.94
14613.41
14752.16
14779.33
14806.50
14852.08
14877.14
14902.19
14939.25
14971.99
15004.73
16426.96
16511.73
16596.50
16598.45
16664.52
16730.58
16995.62
17026.80
17057.98
17104.30
17121.88
17139.47
17139.72
17155.18
17170.65
17176.14
17200.08
17224.03
17226.03
17267.43
17308.84
17341.77
17390.41
17439.05
17440.05
17472.35
17504.66
17568.51
17600.69
17632.87
17772.43
17807.18
17841.94
17852.95
17876.12
17899.29
17969.21
18022.01
18074.81
18226.45
18273.65
18320.85
18433.51
18489.23
18544.96
18549.22
18617.73
18686.25
18687.46
18726.44
18765.41
18766.02
18792.21
18818.40
18819.01
18856.46
18893.91
19049.56
19092.01
19134.45
19492.83
19552.08
19611.34
19882.92
19942.61
20002.29
20474.87
20549.17
20623.46
20711.16
20806.16
20901.17
20902.99
20955.06
21007.13
21008.35
21066.81
21125.28
21126.50
21174.91
21223.33
21224.54
21277.22
21329.90
21331.12
21383.19
21435.26
21651.45
21695.30
21739.15
21739.76
21763.51
21787.26
21787.70
21814.48
21841.25.