Transgender adult reference intervals taking shape

Amy Carpenter Aquino

November 2019—Current sex-specific intervals can be used to interpret hematology results for transgender people using their affirmed gender, say authors of a study published earlier this year.

In a session at the 2019 AACC annual meeting, Dina N. Greene, PhD, DABCC, technical director of Kaiser Permanente Washington laboratories and an author of the study, shared details of the analysis of 172 transgender people recruited between late 2017 and mid-2018 (Greene DN, et al. Clin Chim Acta. 2019;492:84–90). She also reported soon to be published sex hormone reference intervals for transgender adults on stable hormone therapy.

Dr. Greene said she was discouraged from starting the study by people who said the transgender population is too diverse, with the main concern being that some transgender people are on hormone therapy and others are not. Dr. Greene’s response: “People are diverse. We define our population, get a set of reference intervals, and start somewhere. Then we see what else we have to do.”

Dr. Greene and her colleagues collaborated with the University of Iowa Department of Pathology and recruited study participants from two LGBTQ-specific clinics in Seattle and Iowa City. Some of the data were published in Clinica Chimica Acta; other data are in preparation.

Study participants were healthy transgender adults who were at least 18 years old and on stable, gender-affirming hormone therapy for at least one year. “We excluded people if they were diabetics, obese, a current cigarette smoker, had a history of blood clots or cardiovascular disease, if they were HIV positive, had sleep apnea, or had a current pregnancy,” Dr. Greene said.

Researchers tested participants’ whole blood and serum. Measured were complete blood counts, testosterone, estrogen, SHBG, LH, FSH, AMH, progesterone, prolactin, electrolytes, lipids, and HbA1c.

“We collected samples from 79 transgender men and 93 transgender women, ” she said. The average age of study participants was 28.8 years for transgender men and 35.1 years for transgender women. Participants used various modes of hormone administration; injection was the most highly favored method for testosterone—“very common because it’s cheaper,” she said—while more than half of the transgender women chose oral administration for their estrogen. More than half of the transgender women also took an antiandrogen—spironolactone, progesterone, or finasteride—with their estrogen.

The distribution of sex hormone results for people on masculinizing hormones did not closely match cisgender male reference intervals. “The cisgender male intervals say anything greater than 200 ng/dL is normal” for total testosterone levels, Dr. Greene said, while the cisgender male reference range for estradiol is less than 48 pg/mL. If the cisgender reference limit fell outside the 95 percent confidence interval derived from transgender cohorts, the study says, reference change values were used to evaluate if the difference was clinically significant.

“The distribution of results for people on masculinizing hormones for these hormones is different,” Dr. Greene said. The free testosterone measurements, for example, “wouldn’t represent the central 95th percentile; this would represent the central 60th percentile. Therefore, these reference intervals don’t apply.”

Dr. Greene and her collaborators used immunoassays and mass spectrometry to measure sex hormones and concluded that “immunoassay is good enough,” she said. “When do we need mass spectrometry?” is a common question, she added. “For most of the time when you’re measuring this in a general trans male population, the cheap immunoassay is good enough.” And it is not necessary to measure free testosterone. “Total testosterone is generally fine” and will provide the required information.

The authors developed the following sex hormone reference intervals for people on masculinizing hormones: estrogen, less than 100 pg/mL; total testosterone, 180 to 900 ng/dL; and free testosterone, 15 to 170 pg/mL.

For the feminizing cohort, “we have a very similar picture. If you compare their reference intervals to that of cisgender women, they don’t quite apply,” she said.

“With free and total testosterone, it’s similar. There are big discrepancies in the concentration of total testosterone, but this would still be lower than the lower reference limit, so these would kind of group into the same clinical interpretation of, okay, this testosterone is low for this population.”

The authors developed the following sex hormone reference intervals for people on feminizing hormones: estrogen, 30 to 500 pg/mL; total testosterone, less than 200 ng/dL; and free testosterone, less than 20 pg/mL.

They used two immunoassays and looked at the lower limit of both. “For the most part, you’re going to group everything clinically the same whether or not you use mass spec or immunoassay.”

The reference intervals derived for the people on feminizing hormones will be published sooner than those for the masculinizing hormones, Dr. Greene said, adding, “Stay tuned.”

The study of hematology parameters—hemoglobin, hematocrit, and red cell count—for the masculinizing and feminizing cohorts showed good correlation, for the most part, with cisgender male and female reference values, respectively.

The distribution of results for hemoglobin concentration in the masculinizing cohort “fits perfectly, meaning that you can use the cisgender male range,” she said. The commonly used hemoglobin reference interval for cisgender men is 13.0 to 18.0 mg/dL; in the masculinizing cohort, the calculated hemoglobin reference interval was 12.8 to 17.4 mg/dL. “That hormone concentration is what is really driving hemoglobin concentration. This was the same for hematocrit and basically the same for red cell count.”

“If you are a transgender man on stable hormone therapy, your reference intervals should look like cisgender men, which is quite interesting.”

For study participants on feminizing hormones, it “ended up being the exact same story,” Dr. Greene said. “This is also interesting because we know that cisgender women have lower hematological parameters than cisgender men. And we know that testosterone stimulates erythropoiesis, but we also know women menstruate, so it’s been a little vague what the actual physiological mechanism is of the hemoglobin and red cell indices differences.” This showed us, she said, that it is hormone driven. “The cisgender female reference interval fits nicely with this data and is similar for hematocrit and for red cell indices”—a “positive finding,” she added, “because it’s just easy.”

Dr. Greene said she was unable to easily incorporate the reference interval information for transgender patients into the KPWA laboratories’ electronic medical record system, which would accept only a binary gender. She created a workaround that requires appending a comment to the hematocrit test section for every CBC ordered. The comment reads: “For transgender, nonbinary, and gender diverse patients, reference intervals for Hgb, Hct, and RBC can be found here www.KPWAinternallink.com.” (The link will not work outside of KPWA laboratories.)

She created a similar workaround with a comment for the prolactin concentration test. That reference interval has not yet been published.

The comment for race as it relates to estimated glomerular filtration rate provided a precedent for such workarounds. “We already have one example in the lab where we try to include minority populations within the results, even though we don’t have the ability to know someone’s race in our LIS,” she said.

The solution was more complicated for testosterone and estrogen testing. “I couldn’t just append a comment because things are a lot different. So we built four new tests.” The test names are “testosterone for people on masculinizing hormones,” “testosterone for people on feminizing hormones,” “estrogen for people on masculinizing hormones,” and “estrogen for people on feminizing hormones.”

“We didn’t say, ‘for transgender men and transgender women,’ because there are nonbinary people. This was more inclusive.” And the new tests are straightforward, she said. The physician can order testosterone testing for people on masculinizing therapy and see the specific reference levels appended.

“This is helpful if you’re a transgender male, because most of the time you’re not going to suppress your estrogen down low enough that you’re going to look like a cisgender guy,” she said. The new test options present a more realistic picture to patients by showing how well their levels match those of other people who are trying to masculinize. “If the levels are much higher, that’s when you can start to look at aromatization and some other drugs that can be used to try to suppress estrogen.”

Dr. Greene presented the case of a 40-year-old transgender male blood transfusion patient who retained his uterus and ovaries and presented to an outside institution for heavy vaginal bleeding after cervical mass biopsies (Mays JA, et al. Transfusion. 2018;58[3]:823–825). The outside institution reported that the patient’s hemoglobin level was 6 g/dL.

“They transfused him but he kept bleeding,” she said. “They transferred him to the University of Washington and admitted him to the gynecology unit,” where the hemorrhage continued and a massive transfusion protocol was initiated.