Table 2.
General Comparison of the Outputs by FR1 and Leader Primer Sets of the Clinical Implementation Cohorts (MiSeq Platform) Based on the Analysis Strategy Outlined in Figure 3
| Measure | Overall (n = 423) | Leader (n = 218) | FR1 (n = 205) | P |
|---|---|---|---|---|
| Sex, female/male, n (% female) | 154/269 (36) | 75/143 (34) | 79/126 (39) | NS |
| Age, mean (range), years | 63.9 (27–93) | 62.8 (27–90) | 64.9 (36–93) | NS |
| Sample type, n (%) | ||||
| Blood/bone marrow | 348 (82)/45 (11) | 189 (87)/10 (4.5) | 159 (78)/35 (17) | NS |
| FFPE | 27 (6) | 18 (8) | 9 (4) | |
| Other | 3 (1) | 1 (0.5) | 2 (1) | |
| Total reads, mean (range), n | 658,285 (30,958–2,451,851) | 562,757 (32,973–1,294,699) | 759,870 (30,958–2,451,851) | <0.0001 (t-test) |
| Cases with clonal sequences, n (%) | ||||
| 0 | 5/423 (1.2) | 1∗/218 (0.5) | 4/205 (2) | NS |
| 1 | 337/423 (79.7) | 177/218 (81.2) | 160/205 (78) | |
| 2 | 79/423 (18.7) | 39/218 (17.9) | 40/205 (19.5) | |
| 3 | 2/423 (0.5) | 1/218 (0.5) | 1/205 (0.5) | |
| Total clonotypes detected, n | 501 | 258 | 243 | NS |
| Total productive clonotypes, n detected | 435 | 228 | 207 | NS |
| Major productive clonotype, n (%) | ||||
| Mutated (M) | 192/418 (46) | 105/217 (48) | 87/201 (43) | NS |
| Unmutated (UM) | 226/418 (54) | 112/217 (52) | 114/201 (57) | |
| Cases with additional rearrangement(s), n (%)† | ||||
| Productive (P) | 17/81 (21) | 11/40 (27.5) | 6/41 (15) | NS |
| Unproductive (U) | 64/81 (79) | 29/40 (72.5) | 35/41 (85) | |
| M | 34/81 (42) | 19/40 (47.5) | 15/41 (37) | |
| UM | 47/81 (58) | 21/40 (52.5) | 26/41 (63) | |
| Cases with single productive rearrangement, n (%) | ||||
| M | 162/337 (48) | 86/177 (49) | 76/160 (48) | – |
| UM | 175/337 (52) | 91/177 (51) | 84/160 (52) | – |
| Double rearrangement (>1 clonotype), n (%) | 81/423 (19) | 40/218 (18) | 41/205 (20) | |
| P + U | 64/423 (15) | 29/218 (13) | 35/205 (17) | – |
| Mutation status concordant | 59/64 (92) | 28/29 (97) | 31/35 (89) | – |
| P,M + U,M | 20/59 | 12/36 | 8/31 | – |
| P,UM + U,UM | 39/59 | 16/36 | 23/31 | – |
| Mutation status discordant | 5/64 (8) | 1/29 (3) | 4/35 (11) | – |
| P,UM + U,M | 5/5 | 1/1 | 4/4 | – |
| P,M + U,UM | 0 | 0 | 0 | – |
| Double productive (PP) | 15/423 (3.5) | 10/218 (4.6) | 5/205 (2.4) | NS |
| Mutation status concordant | 13/15 | 9/10 | 4/5 | – |
| Both M | 8/13 | 6/9 | 2/4 | – |
| Both UM | 5/13 | 3/9 | 2/4 | – |
| Mutation status discordant | 2/15 | 1/10 | 1/5 | – |
| Triple rearrangement | 2/423 (0.5) | 1/218 (0.5) | 1/205 (0.5) | – |
| Follow-up sample analyzed | 16/81 | 6/40 | 10/41 | – |
| Clonotype, % of total reads (range)‡ | – | |||
| All clonotypes detected | 43 (2.5–92) | 34 (3.5–74.7)∗∗∗ | 52 (2.5–92)∗∗∗ | <0.0001 |
| Major productive | 46 (2.5–92)§ | 37 (2.7–75)∗∗∗§ | 57 (2.5–92)∗∗∗§ | <0.0001 |
| Additional clonotype(s) | 23 (3.2–63)§ | 19 (3.6–63)∗∗§ | 28 (3.2–59)∗∗§ | 0.0026 |
The χ2 test was used to evaluate statistical significance of categorical variable. The t-test was used for continuous variables.
∗∗P ≤ 0.01; ∗∗∗P ≤ 0.0001.
FFPE, formalin-fixed, paraffin-embedded; FR1, framework region 1; NS, not statistically significant.
∗
Single unproductive only.
†
Including case(s) with three clonotypes.
‡
The P values provided in the table reflect a comparison of leader versus FR1.
§
Statistically significant difference seen between percentage of reads of the major and additional clonotypes (P < 0.0001) within both primer sets and overall.