Histologic Analysis of Testes from Prepubertal Patients Treated with Chemotherapy Associates Impaired Germ Cell Counts with Cumulative Doses of Cyclophosphamide, Ifosfamide, Cytarabine, and Asparaginase

Abstract

Cryopreservation of immature testicular tissue is an experimental strategy for the preservation of fertility in prepubertal boys that will be subjected to a gonadotoxic onset, as is the case of oncologic patients. Therefore, the objective of this study was to assess the impact of chemotherapeutic treatments on the testicular histologic phenotype in prepubertal patients. A total of 56 testicular tissue samples from pediatric patients between 0 and 16 years old ( 28with at least one previous chemotherapeutic onset and 28 untreated controls) were histologically analyzed and age-matched compared. At least two 5µm sections from testis per patient separated by a distance of 100 µm were immunostained for the germ cell marker VASA, the spermatogonial markers UTF1, PLZF, UCHL1 and SALL4, the marker for proliferative cells KI67, and the Sertoli cell marker SOX9. The percentage of tubule cross-sections positive for each marker and the number of positive cells per tubule cross-section were determined and association with the cumulative dose received of each chemotherapeutic drug was statistically assessed. Results indicated that alkylating agents cyclophosphamide and ifosfamide, but also the antimetabolite cytarabine and asparaginase were associated with a decreased percentage of positive tubules and a lower number of positive cells per tubule for the analyzed markers. Our results provide new evidences of the potential of chemotherapeutic agents previously considered to have low gonadotoxic effects such as cytarabine and asparaginase to trigger a severe testicular phenotype, hampering the potential success of future fertility restoration in experimental programs of fertility preservation in prepubertal boys.

Testicular Tissue Banking for Fertility Preservation in Young Boys: Which Patients Should Be Included?

“…However, as chemotherapeutic agents are mostly administered in combination, it is difficult to assess the individual risk per agent ( 58 , 67 , 69 ). Moreover, most results are extrapolated from animal or adult studies, making additional research necessary to verify these risk classifications/dosages in children ( 67 , 68 , 75 , 76 , 191 ). Therefore, studies with human-relevant experimental systems such as human organoids ( 18 ) or human organotypic culture combined with xenografting ( 192 ), could be helpful tools to characterize the exact effects of a chemotherapeutic agent on prepubertal testicular tissue.…”

“…Besides this, it remains a challenge to identify the individual risk per molecule, as these are usually administered in combinations ( 58 , 67 , 69 , 74 ). Moreover, most data either come from animal experiments or studies performed in adult men, which makes it difficult to extrapolate these risks to young patients ( 67 , 75 , 76 ). As the exact risk classifications/dosages for children remain unclear, further research on prepubertal exposure is warranted together with regular updates of this classification ( 67 , 68 ).…”

Vinblastine monotherapy induction prior to radiotherapy for patients with intracranial germinoma during the COVID‐19 pandemic

“…In Europe, carboPEI 7,8 is delivered with large volumes of intravenous hydration, can exacerbate pre‐existing DI, particularly where no thirst mechanism is present, and is associated with prolonged inpatient admissions 20 . Such chemotherapy schedules are also associated with short‐ 8 and long‐term 11–13,16 toxicities and, moreover, require central venous access devices for delivery, with associated infection and thrombosis risk 14,15 . A further UK intracranial germinoma case, in addition to the two formally described here, received two vinblastine doses to complete induction as a “bridge” to radiotherapy with stable radiological appearances; this patient developed ifosfamide encephalopathy 23 during standard‐of‐care carboPEI chemotherapy and experienced a fall and subdural hematoma.…”

“…In addition to the challenges of managing DI, carboPEI chemotherapy is associated with short‐term toxicities of myelosuppression, vomiting and/or diarrhoea, electrolyte disturbances, which may lead to seizures, renal impairment and elevation of liver enzymes 8 . Long‐term sequelae of these drugs include ototoxicity from cisplatin 11 and reduced fertility from alkylating agents (ifosfamide) 12,13 . Current chemotherapy regimens also require the use of indwelling central venous access devices, which are associated with increased risk of infection 14 and thrombosis 15 and which may affect quality of life.…”

Achievement of complete in vitro spermatogenesis in testicular tissues from prepubertal mice exposed to mono- or polychemotherapy

“…The majority of prepubertal patients have received chemotherapy regimens, considered to be at low gonadotoxic risk, before the preservation of their testicular tissues 22 , 23 . Some studies have demonstrated that the administration of chemotherapy before puberty led to a decrease in the number of spermatogonia, especially when alkylating agents were included in the therapy regimen 24 – 26 . The depletion of the spermatogonial stem cell (SSC) pool could compromise the chances of fertility restoration.…”