Lawyers for Adam Rosendorff, the former Theranos Inc. lab director who testified against founder Elizabeth Holmes in her criminal-fraud trial, asked a judge Wednesday to reject a subpoena from Ms. Holmes as she seeks a new trial.

Dr. Rosendorff emerged as a central witness for the prosecution in a monthslong trial of Ms. Holmes, who was convicted in January on four counts of wire fraud and conspiracy for misleading investors about the blood-testing startup’s technology.

Last month, Ms. Holmes asked a federal judge for a new trial, citing a visit to her California home by Dr. Rosendorff, in which he allegedly expressed regret for his role in her prosecution, according to a court filing.

Dr. Rosendorff’s lawyers dispute that account, saying in the filing Wednesday that he doesn’t recall making those statements and that they don’t accurately reflect how he felt then or now.

“He wanted to be able to forgive her for the pain and suffering her actions have caused in his life,” his lawyers wrote in the filing. “He wanted to be able to express his condolences that her child may grow up without a mother” if Ms. Holmes receives a lengthy prison sentence, they wrote. “And he wanted to be able to draw a line under the nine-year saga of his employment at Theranos, the harassment and intimidation directed at him after he left, his participation in the subsequent investigation, and his high-profile testimony in the trials in this case.”

Dr. Rosendorff’s lawyers said he showed up at Ms. Holmes’s house on Aug. 8 after spending the afternoon in Palo Alto and visiting the site where Theranos’s building once stood. It had been torn down and a residential complex was built in its place, according to the filing, making the site unrecognizable.

Adam Rosendorff, left, was a central witness for the prosecution during Elizabeth Holmes’s trial.

Photo: David Paul Morris/Bloomberg News

Dr. Rosendorff wanted to move on, and felt like he needed to have a conversation with Ms. Holmes to gain closure, the filing says. That led to a phone call to her lawyer and then an unannounced visit to her home.

During the August visit, Dr. Rosendorff didn’t speak directly to Ms. Holmes but to her partner, Billy Evans, who answered the door, according to Mr. Evans’s account of the exchange, which was filed into court record.

After the visit, Ms. Holmes asked a federal judge for a new trial, saying the former lab director told her partner that he “feels guilty,” “that the government made things sound worse than they were” and that “he felt like he had done something wrong,” according to court filings. In a subsequent subpoena, Ms. Holmes’s lawyers sought emails and communications from the former lab director regarding trial testimony and the prosecution team.

Judge Edward Davila, who oversaw the trial of Ms. Holmes, granted an evidentiary hearing set for Oct. 17 to ask Dr. Rosendorff if his testimony during the trial was truthful and if he felt manipulated by the government.

Dr. Rosendorff’s lawyers say the subpoena is oppressive and unreasonable given the scope of the hearing next week. They also cite a declaration made by Dr. Rosendorff on Sept. 15 in response to Ms. Holmes’s request for a new trial, in which he said he answered questions truthfully and had no reason to believe the government misrepresented Ms. Holmes.

Ms. Holmes was acquitted of several counts of defrauding patients, the part of the trial for which Dr. Rosendorff’s testimony was most relevant.

Her sentencing date, which was previously scheduled for Oct. 17, has been moved to Nov. 18.

Write to Meghan Bobrowsky at Meghan.Bobrowsky@wsj.com