High Prevalence of Clonal Hematopoiesis in Patients Undergoing Hip Replacement Surgery - Implications for Experimental Models and Association with Autoimmune Disease

Background:

Clonal hematopoiesis (CH) is characterized by the presence of genetic alterations in the peripheral blood (PB) of individuals without clinical manifestation of a hematologic malignancy. Diverse cohorts have been screened for CH and it was observed that CH increases progressively with age.

Prosthetic hip replacement is among the most common surgical procedures in aging persons. Bone marrow (BM) from femoral heads obtained during hip replacement surgery provides an opportunity to genetically and functionally characterize CH in the marrow compartment.

Aims:

We prospectively studied CH in patients (pts) undergoing hip replacement surgery, aiming to characterize the spectrum of CH in this population, understand associations between CH and clinical parameters, longitudinally monitor CH over time, and acquire CH BM specimens for functional analyses.

Methods:

The data presented here includes 159 pts undergoing hip replacement surgery, with a median age of 72 years (range, 18-86 yrs). PB samples (n=56) and/or BM samples obtained from femoral heads (n=116 samples from 113 pts) were screened for CH-associated variants in 68 genes using a targeted sequencing approach (variant allele frequency [VAF] cut off, 1%; Rothenberg-Thurley et al., Leukemia 2018). Follow-up PB samples were available for 33 pts, 6-18 months after surgery.

Results:

We achieved a median sequencing coverage of 904x (interquartile range, 798-990x). In 65/159 pts (41%), we detected CH with VAFs of >2%, and CH with low mutation burden (VAF 1-2%) was identified in an additional 17 pts. In sum, CH was present in 82/159 pts (52%) with VAFs ranging from 1.0-31.7% (median, 2.8%). CH became progressively more frequent with aging (50-59y: 29%, 60-69y: 38%, 70-79y: 59% and 80-89y: 83%, Figure 1A). In addition, the number of variants per pt increased with age (p=0.02), while the VAF of individual variants showed no association with age. Compared to pts without CH, pts positive for CH had lower hemoglobin levels (median, 13.7 vs 12.7g/dl; p=0.02) and a higher MCV (median, 89 vs 92fl; p=0.03). Interestingly, CH was also significantly associated with presence of autoimmune disease as CH was detected in 10/12 (83%) of pts with autoimmune disease compared to 72/147 (49%) of pts without (p<0.001). Furthermore, CH was also enriched in pts with prior history of malignancy (12/18; 67% vs 70/141; 50%; p<0.01).

Most variants were detected in DNMT3A (48 pts, 30%), TET2 (25 pts, 16%) and ASXL1 (6 pts, 4%). Notably, we observed a shifting mutation spectrum with age. While younger pts mostly harbored variants in DNMT3A, the proportion of pts with TET2 variants increased with age (Figure 1B).

For 10 pts, we collected both PB and BM at the time point of surgery. The mutation profile of the two samples was concordant for each pt; however, VAFs were significantly higher in the BM compared to matched PB samples (median VAFs, 4.2% vs 3.4%, p=0.01).

In 3 pts with CH, both femoral heads were removed simultaneously, enabling pairwise variant analysis. Two pts showed identical mutation patterns in BM obtained from the right and left femoral heads, with little differences in VAFs (DNMT3A; VAF 5.1%/4.7% and NOTCH1; VAF 2.0%/1.4%). In contrast, in another pt, two variants were detected in both sides (JAK2; VAF 6.0%/6.7% and DNMT3A; VAF 0.9%/1.1%) but one variant was present only in one BM sample (ASXL1; VAF 1.3%), suggesting the possibility of spatial heterogeneity of CH clones in the marrow compartment.

Follow-up samples 12 months after screening were available for 16 pts without CH. Gain of variants over this period was not observed in these samples. In the group of pts with CH (n=17), follow-up samples were available after 6 months (n=16), 12 months (n=12) and 18 months (n=3). Of the 24 variants identified at screening, all but one (RUNX1; VAF 2.7%) were still detectable in the sequential PB samples. Overall, VAFs remained relatively stable over time (Fig. 1C).

Conclusions:

We screened a total of 159 pts undergoing hip replacement and observed an unexpectedly high CH prevalence of 52%. In light of this data, the routine use of femoral heads as a source of healthy control BM in experimental setups should be reconsidered. Analysis of follow-up samples indicates stability of the observed mutation patterns over time. Our data additionally suggest an interplay between CH and autoimmune disease which warrants further investigation.

Author contributions:

LH and JH contributed equally to this work