Myelotoxicity of Peptide Receptor Radionuclide Therapy of Neuroendocrine Tumors: A Decade of Experience

Abstract

Aim:

This review of the literature, and the authors' own decade of experience with lutetium-177-octreotate-capecitabine±temozolomide peptide receptor radionuclide therapy (PRRT)-chemotherapy of GEPNETs, analyses the risk of both short- and long-term hematotoxicity.

Background:

Myelodysplastic syndrome (MDS) and acute leukemia (AL) have been associated with PRRT in heavily pretreated patients with a history of exposure to alkylating agents. Commenced 15 years ago, PRRT is now becoming established as first- and second-line therapy for gastroentero pancreatic neuroendocrine tumors (GEPNETs), and early treatment minimizes myelotoxicity, which is the most significant potential adverse event following PRRT.

Results:

Sixteen key articles involving primary research were identified. A total of 2225 patients were treated (2104 treated with PRRT monotherapy and 121 with PRRT combined with chemotherapy). The average age of patients in these studies ranged from 53 to 64 years with median duration of follow-up ranging from 6 to 62 months. Short-term myelotoxicity was observed in 221 patients (10%), occurring in 213 of 2104 patients treated with PRRT monotherapy and 8 of 121 patients treated with PRRT combined with chemotherapy. Acute toxicity manifested as modest self-limited grade 3/4 toxicity (CTCAE or WHO), most often affecting platelets during the first cycle of treatment. Toxicity manifesting early was easily managed with dose modification or therapy cessation and was ameliorated by appropriate patient selection. MDS/AL was a rare stochastic event occurring in 32 (1.4%) patients. Where bone marrow biopsy was performed, cases of MDS displayed cytogenetic abnormalities, consistent with secondary MDS. Factors associated with myelotoxicity included age >70 years, impaired renal function, baseline cytopenias, prior number of therapies, prior chemotherapy (alkylating agents), and prior radiotherapy.

Conclusion:

Early therapy with PRRT-containing regimens improves outcomes, minimizes myelotoxicity, and renders the risk of MDS and AL negligible.

Background

Targeted tumor therapy of neuroendocrine tumors (NETs) using somatostatin analog peptide receptor radionuclide therapy (PRRT) was introduced 15 years ago using ¹¹¹In, ⁹⁰Y, and ¹⁷⁷Lu-Octreotide.^1

–4

The advent of combination radionuclides or radiosensitizing concomitant chemotherapy has improved the overall response rate (ORR) and progression-free survival (PFS) in comparison with radiopeptide monotherapy,^5
–7 and this review compares the myelotoxicity of these novel approaches. In the last decade, PRRT has been extensively researched and is now considered an early treatment option in the setting of inoperable, well-differentiated, somatostatin receptor-positive metastatic gastroentero pancreatic NET (GEPNET).⁸ Furthermore, there is increasing evidence of efficacy for ¹⁷⁷Lu-octreotate-based salvage therapies for progressive advanced GEPNET.⁹

The nonhematologic toxicities of PRRT have been largely obviated by procedural modifications such as renal protection by amino acid infusion or gelofusine and optimization of antiemetic regimens to avoid nausea.^8,10,11

Despite the large body of evidence for safety and efficacy, and now preliminary results from the multicenter registrational Phase III randomized controlled trial (NETTER 1), PRRT for GEPNETs is still considered “investigational” with particular concerns regarding myelotoxicity. The long-term risks of myelodysplastic syndrome (MDS) and acute leukemia (AL) have been raised and are reflected in the 2016 ENETS consensus guidelines.¹²

Myelotoxicity is now deemed to be the most significant potential adverse event following PRRT, and this review of the literature, complemented by the authors' own decade of experience with ¹⁷⁷Lu-octreotate-capecitabine-temozolamide PRRT-chemotherapy of GEPNETs, analyses the risk of hematologic toxicity as manifested by thrombocytopenia and neutropenia, in both the short- and long-term follow-up, and also documents the incidence of MDS and AL.

Method

All of the literature on myelotoxicity in association with PRRT of NETS reported in articles published over the past 15 years has been reviewed and compared with the authors' own experience of lutetium-177 PRRT combined with octreotate-capecitabine-temozolomide radiosensitizing chemotherapy. Relevant primary research concerning myelotoxicity and PRRT was identified using medical databases. Key articles published from 2000 onward were obtained primarily from PubMed, Ovid, and Cochrane databases. Specific focus was given to articles published in Cancer Biotherapy and Radiopharmaceuticals, European Journal of Nuclear Medicine and Molecular imaging, Neuroendocrinology, Journal of Clinical Oncology and Endocrine Related Cancer.

Results

Sixteen key articles involving primary research were identified (Table 1); 12 articles relating to PRRT as monotherapy and 4 articles relating to PRRT in combination with chemotherapy and/or novel agents. A total of 2225 patients were treated (2104 treated with PRRT monotherapy and 121 with PRRT combination), all of whom had been exposed to prior therapies, including somatostatin analogs, alkylating agents, radiotherapy, prior PRRT, and surgery, with the majority having had exposure to greater than two prior lines of treatment. The average age of patients in these studies ranged from 53 to 64 years with median duration of follow-up ranging from 6 to 62 months. Short-term myelotoxicity was observed in 221 patients (10%), occurring in 213 of 2104 patients treated with PRRT monotherapy and 8 of 121 patients treated with PRRT combination. Acute toxicity manifested as modest self-limited grade 3/4 toxicity (CTCAE or WHO) most often affecting platelets, then white blood cells (WBC), and finally hemoglobin, and was commonly observed during the first cycle of treatment, with the lowest nadir predictive of time taken for recovery. Toxicity manifesting early was easily managed with dose modification or therapy cessation and was ameliorated by appropriate patient selection based on age, prior therapies, comorbidities, and adequate baseline myeloid function.

Table 1.

Summary of Reported Myelotoxicity of Peptide Receptor Radionuclide Therapy for Neuroendocrine Tumor

Author	Number of patients included in analysis	Average age (years)	Prior therapy	PRRT treatment	G 3/4 hematologic toxicity (%)	Median f/up (m)	MDS/AL (%)
PRRT monotherapy
Kwekkeboom et al.,	504	59	Surgery/SSA/chemo/radio	177-LuDOTATATE	48 (9.5%)	19	4
Bodei et al.¹⁹	51	57 (median)	SSA	177-LuDOTATATE	1 (1.9%)	29	Nil
Pfeifer et al.¹⁴	69	58	SSA/chemotherapy/IFN		17 (25%)	—	1 (1.4%)
			n = 53	90-YDOTATOC
			n = 12	90-YDOTATOC +177-LuDOTATATE
			n = 4	177-LuDOTATATE
Sansovini et al.²⁰	52	61	SSA/90-Y/chemo/radio	177-LuDOTATATE	0	29	Nil
Sabet et al.²¹	33	61	PRRT with initial response	177-LuDOTATAE	7 (21%)	37	Nil
Delpassand et al.²³	32	63.4	SSA/chemo	177-LuDOTATATE	4 (12.5%)	14 (mean)	Nil
Paganelli et al.²²	43	64 (median)	SSA/surgery/chemo	177-LuDOTATATE	0	38	Nil
Sabet et al.²⁴	61	62	SSA/chemo/surg/radio	177-LuDOTATATE	5 (8%)	62	Nil
Bodei et al.¹⁵	807	53	SSA/chemo/surg/radio		77 (9.5%)	30	21 (2.6%; 13 MDS, 6 MDS t/f to AL, 2 AL)
			n = 360	90-YDOTATOC	51 (14.2%)
			n = 157	90-YDOTATOC +177-LuDOTATATE	17 (10.8%)
			n = 290	177-LuDOTATATE	9 (3.1%)
Bergsma et al.²⁵	320	—	Chemo/radio (22%)	177-LuDOTATATE	34 (11%)	—	Incidence not reported
Mariniello et al.¹⁶	112	60.3	Not specified		7 (6.25%)	45.1	0
			n = 44	90-YDOTATOC	5 (11.4%)
			n = 21	90-YDOTATOC +177-LuDOTATATE	2 (9.5%)
			n = 47	177-LuDOTATATE	0
Brieau et al.²⁶	20		PRRT/chemo/radio	177-LuDOTATATE	6 (30%)	37	4 (20%)
PRRT in combination
van Essen et al.³⁰	7	62 (median)	No prior chemo/PRRT	177-LuDOTATATE+capecitabine	2 (28%)	—	0
Claringbold et al.⁵	33	60	SSA/surgery/chemo	177-LuDOTATATE+capecitabine	1 (3%)	16	0
Kesavan et al.³²	65	60	SSA/chemo/surg/radio		9 (13.8%; 1 ST, 8 LT)		2 (5.4%)
			n = 28	177-LuDOTATATE+capecitabine	3 (10.7%; 1ST, 2 LT)	60	0
			n = 37	177-LuDOTATATE+capecitabine+temozolomide	6 (16.2%; 0 ST, 6 LT)	36	2 (5.4%)
Claringbold et al.⁵	16	63 (median)	SSA/chemo/surg/PRRT		3 (18.75%)	6	0
			n = 4	177-LuDOTATATE+everolimus 5 mg
			n = 9	177-LuDOTATATE+everolimus 7.5 mg
			n = 3	177-LuDOTATATE+everolimus 10 mg

AL, acute leukemia; chemo, chemotherapy; LT, Long-term; MDS, myelodysplastic syndrome; pt, ; PRRT, peptide receptor radionuclide therapy; radio, Irradiation; SSA, somatostatin analog; ST, short-term; surg, surgery; t/f, transformed.

Long-term toxicity in the form of MDS and AL was a rare stochastic event occurring in only 32 (1.4%) of all the patients treated with PRRT (24 cases of MDS, 6 cases of MDS with subsequent transformation to AL, and 2 cases of AL alone). Where bone marrow biopsy was performed, the majority of cases of MDS displayed complex cytogenetic abnormalities, consistent with secondary MDS following exposure to chemotherapy with alkylating agents or irradiation. The development of MDS was a risk factor for AL. No significant difference in relative risk of MDS/AL was noted between patients salvaged with PRRT monotherapy or PRRT combination (1.4% and 1.6%, respectively).

Factors associated with development of significant short- and long-term myelotoxicity included age >70 years, impaired renal function, baseline cytopenias, prior number of therapies, prior chemotherapy (especially alkylating agents), and prior radiation therapy. The latter two factors were associated with a significant development of MDS and AL.

Toxicity of PRRT as monotherapy

Indium-111-octreotide

Indium-111-octreotide PRRT achieved only modest tumor regression and ORR, at the cost of considerable myelotoxicity, including the development of MDS and AL.^13,14 In their Rotterdam cohort, Valkema et al. treated 50 heavily pretreated patients with ¹¹¹In-Octreotide with administered activities ranging from 20 to 160 GBq. Three out of six patients receiving greater than 100 GBq of administered activity developed MDS/AL, thus defining the maximum tolerated dose (MTD). Additional risk factors for myelotoxicity included bone marrow involvement and previous exposure to radiotherapy.¹³ In a similar study from New Orleans, Anthony et al. treated 27 patients with ¹¹¹In-Octreotide at monthly administered activities of 6.6 GBq. Once again patients were heavily pretreated and Octreoscan^® positive at enrollment. Grade 3/4 toxicity affecting platelets and neutrophils was observed in four patients (15%), occurring predominantly after two cycles of therapy. No MDS/AL was observed.¹⁴ This toxicity directly relates to the high marrow exposure to the Auger electron emission of ¹¹¹In PRRT.⁴

Yttrium-90-DOTATOC

In a 2004 review, Bodei et al. published their Milan experience,¹⁰ which noted reversible grade 3/4 toxicity of WBC and/or platelets in three out of seven (43%) patients who received an administered activity of 5.18 GBq, defining it as the MTD. In addition, 31 out of 40 (77.5%) of these treated patients developed grade 3/4 lymphocyte toxicity. Development of MDS/AL was not reported. This incidence of toxicity was, in part, attributed to the longer range (up to 10 mm) of the relatively high-energy (E_max 2.27 MeV) β emissions of ⁹⁰Y and increased bone marrow irradiation contributing both to the crossfire and bystander effects.¹⁰ Furthermore, the lymphocyte toxicity observed is likely to also relate to in vivo saturation of normal splenic tissue, which acts as a reservoir for WBC, thus potentiating toxicity.

In 2011, Pfeifer et al. published early Swiss experience of myelotoxicity¹⁵ with Yttrium-90 DOTATOC, either as monotherapy or in combination with Lutetium-177-DOTATATE. Of the 69 patients studied, 53 had ⁹⁰Y-based therapy, 12 ⁹⁰Y+¹⁷⁷Lu, and 4 ¹⁷⁷Lu alone. Of the 69 patients, 17 (25%) developed grade 3+ toxicity (6 cases of anemia required red cell support in the ⁹⁰Y-DOTATOC monotherapy group, 5 cases of neutropenia in the ⁹⁰Y+¹⁷⁷Lu combination group, and 6 of thrombocytopenia in the ¹⁷⁷Lu-DOTATATE monotherapy) with 1 patient (1.4%) from the ⁹⁰Y monotherapy cohort going on to develop MDS. Although no comment was offered on associations, it is evident, given the distribution of patients, that exposure to ⁹⁰Y plays an important role in the development of myelotoxicity.

The Milan group recently published their long-term PRRT data in 807 NET patients,¹⁶ demonstrating the increased incidence of hematotoxicity in patients receiving ⁹⁰Y-DOTATOC as monotherapy when compared with Lutetium-177-DOTATATE alone, or in combination with ⁹⁰Y-DOTATOC.¹⁶ Median administered activities were 10 GBq with a median number of four cycles and 6.4/12.7 GBq over a median five cycles for ⁹⁰Y-DOTATOC monotherapy and ¹⁷⁷Lu+⁹⁰Y combination therapy, respectively. Fifty-one of the 360 patients (14.2%) who received ⁹⁰Y-DOTATOC monotherapy developed grade 3/4 toxicity of WBC and/or platelets. This was markedly increased in comparison with the ¹¹⁷Lu+⁹⁰Y combination group, of which 17 out of 157 patients (10.8%) developed grade 3/4 toxicity, compared with only 9 out of 290 patients (3.1%) in the ¹⁷⁷Lu-DOTATATE monotherapy group (none of whom developed a grade 4 toxicity). The presence or subsequent development of renal dysfunction was strongly associated with development of acute myelotoxicity. Nineteen of 807 patients (2.35%) developed MDS, with 8 (1.1%) of these patients transforming into AL. The major risk factors for development of MDS/AL included prior chemotherapy, bone marrow compromise (NET involvement and/or prior radiotherapy), and the severity of platelet toxicity. Of the eight AL cases, one was promyelocytic leukemia, three myeloid, three lymphoblastic, and one unspecified. As acknowledged by the authors, MDS most commonly transforms to a myeloid leukemia, while promyelocytic and lymphoid leukemias are commonly de novo, often presenting with distinct pathogenic genetic aberrations not commonly associated with prior alkylating therapies. Thus, the incidence of AL in this study cannot be solely attributed to PRRT. Furthermore, no patient receiving ¹⁷⁷Lu-DOTATATE monotherapy developed AL.¹⁶ Interestingly, individuals who developed persistent toxicity had a significantly less cycles of PRRT (mean 387 days vs. 658 days for transient toxicity, p < 0.004), suggesting that additive toxicity in pretreated patients manifests early, acting as a potential warning sign.¹⁶

Mariniello et al. also noted increased toxicity of ⁹⁰Y-DOTATOC in the Milan group's recent review of PRRT for bronchopulmonary carcinoid.¹⁷ Of the 112 patients (44 receiving ⁹⁰Y monotherapy, 21 ⁹⁰Y+¹⁷⁷Lu, and 47 ¹⁷⁷Lu monotherapy), all 7 who experienced grade 3/4 hematotoxicity were exposed to ⁹⁰Y. Furthermore, the majority of these (5/7) were in the Yttrium-90-DOTATOC monotherapy group. No case of MDS/AL was reported. The study concluded that Lutetium-177-DOTATATE is the best available option for PRRT of metastatic progressive bronchial NET and advocated its use at an earlier stage of the disease.¹⁷

Retreatment with Yttrium-90 PRRT

Machta et al. recently presented the London experience of PRRT retreatment of metastatic NETs.¹⁸ The 48 patients included in their review had all received two or more cycles of PRRT with 46 of the 48 having received first-line ⁹⁰Y-PRRT with 10% incidence of bone marrow toxicity. These patients were subsequently retreated >12 months later with either ⁹⁰Y-DOTATATE (30 of 48 patients) at an administered activity of 3.2 GBq/cycle or ¹⁷⁷Lu-DOTATATE at a standard administered activity of 7.4 GBq/cycle. The median time to PRRT retreatment was 27 months. In the PRRT retreatment cohort, bone marrow toxicity was observed in 10 patients (21%), with the majority experiencing prolonged suppression. This finding was coupled with a 76% disease control rate (DCR), leading the authors to conclude that in the setting of progressive disease, the risk of PRRT retreatment is outweighed by its benefit.¹⁸ Despite these findings, recent data, as previously described, clearly demonstrate increased efficacy and safety with ¹⁷⁷Lu-DOTATATE in comparison to ⁹⁰Y-PRRT for NET, and as such, much of the international experience has been centered upon this approach.

Lutetium-177-DOTATATE

In 2008, the Rotterdam experience of Kwekkeboom et al. in 504 patients (458 GEPNETs)¹⁹ comprised of three cohorts, 7 patients treated at 3.7 GBq, 16 patients at 5.6 GBq, and the remainder at 7.4 GBq. Of the 504 patients treated, 47 (9.5%), developed WHO grade 3/4 hematologic toxicity, with 4 confirmed cases of MDS 2–3 years after completion of PRRT. With regard to the serious complication of MDS, the authors concluded that prior alkylating therapy was an important predisposing factor with the majority of cases having had prior exposure.¹⁹

The Milan Phase I–II study²⁰ treated 51 consecutive patients with 177-Lu-DOTATATE in two separate groups at escalating activities: 3.7–5.1 and 5.1–7.4 GBq, respectively. Patients of median age 57 years had a variety of somatostatin receptor two (SSR2)-positive metastatic NETs with prominent liver involvement (41/51). Eighteen patients had “extended” disease with heavy infiltration of the liver, bone marrow, and lymph nodes. One patient experienced grade 3 toxicity (thrombocytopenia and leukopenia). Toxicity appeared to be most prevalent during cycles 3 and 4, the interval between cycles being 6–9 weeks. Over the 24 months of follow-up, no significant long-term hematotoxicity was noted, with most patients returning to within 10% of baseline with respect to platelet count and hemoglobin, and to within 20% of baseline for WBC. No clinically significant sequelae of these cytopenias were remarked. The MTD was not reached. Objective response (OR) was seen in 15/46 (32.6%) of assessable patients, with a median time to progression of 36 months.²⁰

Sansovini et al. in their 2013 study from Meldola²¹ applied the by now well-established safe standard administered activity, 7.4 GBq ¹⁷⁷Lu-DOTATATE, but reduced it to 4.6 GBq in the cohort of their patients with higher risk factors for renal impairment and myelotoxicity. Of the 52 heavily pretreated patients (26 in each cohort), the cumulative activity across five cycles was 27.8 and 18.5 GBq. No significant acute or long-term hematotoxicity was observed over the 2-year median follow-up period. The 27.8 GBq cohort failed to reach median PFS, whereas it was attained at 20 months for the 18.5 GBq group. There was no difference in myelotoxicity between the two groups.²¹

Sabet et al. in their 2014 retrospective study from Bonn,²² reviewed outcomes of ¹⁷⁷Lu-DOTATATE salvage in 33 patients with progressive metastatic GEPNETs who had demonstrated a prior response to PRRT. Administering a mean cumulative activity of 44.3 GBq across one to four cycles, 21% (7/33) experienced grade 3/4 hematotoxicity, predominantly affecting WBC and platelet counts. No cases of MDS/AL were recorded over the median follow-up of 36 months. The higher than expected incidence of hematotoxicity illustrates the potential for myeloid damage with repeat exposure to radiation. This risk may be deemed acceptable, given that improved disease control was achieved in 67% of this relapsed population. In this study, there may have been insufficient time for clonal evolution, and the lack of MDS/AL may simply reflect the relatively short period of follow-up. The authors acknowledged this, but made the observation that these prognostically limited patients should not be precluded from salvage PRRT on the basis of an unsubstantiated potential risk of late side-effects.²²

Paganelli et al. from Meldola adopted a similar approach,²³ in a Phase II study, prescribing the administered activity according to hematologic and or renal risk. Forty-three patients were assessed (25 full dose and 18 reduced dose), and no significant hematotoxicity was observed over the 3+ years of follow-up. Although the overall response rate (ORR) of 7% is minimal, this trial highlighted a DCR of 84%, with the majority of patients maintaining stable disease, thus demonstrating efficacy for both levels of administered activity, while demonstrating the safety of this approach in “at-risk” patients.²³

The first American Phase II experience of PRRT with ¹⁷⁷Lu-DOTATATE was a Houston-based single-center trial by Delpassand et al.²⁴ Of the 32 evaluable patients, 5 received one cycle at an average administered activity of 7.33 GBq, 8 patients received two cycles at an average administered activity of 14.7 GBq, 5 patients received three cycles at an average administered activity of 21.8 GBq, and 19 patients received four cycles at an average administered activity of 28.7 GBq. 12.5% (4/32) experienced reversible grade 3 hematotoxicity, returning to baseline within 18 weeks. No episode of grade 4 toxicity was recorded. No patients required transfusion support. A significant correlation between prior (unspecified) chemotherapy and grade 2/3 toxicity was noted (p-value = 0.036). Increased grades of toxicity were noted in patients receiving greater than two cycles of therapy. Finally, in addition to a promising ORR of 31% and median PFS of 16.1 months, they noted an impressive improvement in Karnofsky Performance Scores after treatment with ¹⁷⁷Lu-DOTATATE.²⁴

Following the Bonn experience, Sabet et al.²⁵ in 2015 demonstrated the safety and efficacy of ¹⁷⁷Lu-DOTATATE in advanced midgut NET. Sixty-one patients were treated at a higher activity of 7.9 GBq for each of four cycles, followed up over a 4-year period. Five of 61 patients (8.2%) experienced grade 3 hematotoxicity, which was reversible, with recovery of counts occurring within 17 months of PRRT completion. Graduated lineage-specific toxicity was noted 3–10 weeks after at least one cycle of therapy, with sequential cycles of therapy preferentially affecting hemoglobin (two cycles), platelet (three cycles), and WBC counts (seven cycles). All patient blood counts normalized following cessation of therapy, with an average time to normalization of counts of 17 months post-therapy. No case of MDS/AL was reported. This minimal self-limited hematologic toxicity was accompanied by an impressive DCR of 91.8% and median PFS of 33 months.²⁵

In 2014, the Bangalore group of Danthala et al.⁹ published their 5-year experience of ¹⁷⁷Lu-DOTATATE PRRT for NETs. Although the specific incidence and the grades of toxicity were not documented, of the six fatalities observed, none was attributed to hematotoxicity or related sequelae. In this clinical study, they demonstrated a marked outcome advantage in patients receiving greater than two cycles of PRRT with a mean PFS of 45.6 months compared with 8.3 months in those treated with one to two cycles.⁹

The previously cited Milan review of 807¹⁶ patients included 290 who received ¹⁷⁷Lu-DOTATATE-based therapies (278 monotherapy, 12 in combination with capecitabine 1500 g/day).¹⁶ Only 9 of these 290 patients developed grade 3/4 toxicity of WBC and/or platelets. The ¹⁷⁷Lu-DOTATATE group was less likely to develop MDS, and those patients who did were less likely to develop AL.¹⁶ Indeed, practice changed over the period of this study, and encouraging results of PRRT led to not only its first-line use but also a switch of radionuclide from ⁹⁰Y to ¹⁷⁷Lu. Only 28.6% of the ¹⁷⁷Lu-DOTATATE monotherapy group had had prior exposure to chemotherapy and only 3.1% had prior radiotherapy, and myelotoxicity was insignificant in this relatively treatment-naive group.¹⁶ Mariniello et al. observed similar safety data for ¹⁷⁷Lu-DOTATATE in their review of PRRT for bronchopulmonary carcinoid, in which no significant hematotoxicity was noted in patients receiving ¹⁷⁷Lu PRRT monotherapy, fractioned so as to not exceed the renal dose threshold of 25–27 or 30–40 Gy of the biologically effective dose.¹⁷

Bergsma et al. in 2015 reviewed the Dutch experience of 320 patients treated between 2000 and 2007.²⁶ Thirty-four patients (11%) experienced grade 3/4 toxicity; thrombocytopenia in 25, leukocytopenia in 17, anemia in 10, and pancytopenia in 3. Peak acute myelotoxicity occurred at 4–8 weeks after PRRT administration in these patients. Thirty patients experienced chronic toxicity lasting for >6 months, or requiring transfusion support. The development of grade 3/4 toxicity was associated with prior WBC ≤4 × 10⁹/L, age more than 70 years, and extensive tumor mass with high uptake on ¹¹¹In-octreotide γ imaging (OctreoScan^®). A greater than 15% reduction in hemoglobin after cycle 1 was associated with prior exposure to radiotherapy, suggesting some depletion of myeloid reserve due to previous treatment. Previous chemotherapy was not associated with an early risk of hematotoxicity. Count reductions of ≥25% from baseline after first therapy were, however, observed in patients with baseline renal impairment. MDS and AL were recorded, but Bergsma et al. noted these as “rare complex stochastic events” and plan to report them as a separate study.²⁶ Given that the majority of patients in this 2015 report received 7.4 GBq ¹⁷⁷Lu-DOTATATE, without adjustment for baseline parameters, at-risk groups (age >70 years, almost 20% of the cohort), renal impairment, or a history of significant exposure to prior chemotherapy, especially with alkylating agents, the resulting relatively high incidence of hematotoxicity is to be anticipated. Dose modification of PRRT, accepting the concomitant reduction in efficacy, may be advisable in such patients to minimize the myelotoxic risk.

Brieau et al.²⁷ in their French study reported a very high incidence of hematotoxicity and MDS/AL in their series of 20 GEPNET patients salvaged with ¹⁷⁷Lu-DOTATATE. Six of the 20 patients (30%) developed grade 3/4 toxicity, predominantly of neutrophils and platelets (one patient grade 3 neutropenia during treatment, two patients grade 3 thrombocytopenia at 12 and 18 months, respectively, one patient grade 3 neutropenia and grade 4 thrombocytopenia at 12 months, one patient grade 3 neutropenia and thrombocytopenia, and one patient developed grade 4 pancytopenia). Of the five patients with long-term toxicity, three went on to develop MDS and one AL. This 20% incidence was clearly linked to the number of prior therapies, exposure to alkylating agents, bone metastatic burden, and the development of early hematologic toxicity.

Strosberg et al.²⁸ recently presented the U.S. data on ¹⁷⁷Lu-DOTATATE efficacy and safety from their NETTER-1 Phase III trial. Two hundred thirty patients randomized to either ¹⁷⁷Lu-DOTATATE at standard administered activity of 7.4 GBq for four cycles or long acting Octreotide 60 mg every 4 weeks. Only six patients (5%) in the ¹⁷⁷Lu-DOTATATE group experienced dose modifying toxicity with grade 3 or 4 neutropenia occurring in 1%, thrombocytopenia in 2%, and lymphopenia in 9%. The median PFS was not reached in the ¹⁷⁷Lu-DOTATATE therapy cohort in comparison to 8.4 months for the long acting Octreotide cohort. Thus, the preliminary data from this Phase III study showed a potentially clinically meaningful and significantly increased PFS, and OR, demonstrated by ¹⁷⁷Lu-DOTATATE for GEPNET patients, with the suggestion of an overall survival (OS) advantage, without significant hematotoxicity.²⁸

Sansovini et al.²⁹ updated the Meldola experience with presentation of their long-term follow-up data for GEPNET patients treated with ¹⁷⁷Lu-DOTATATE PRRT. Of the 65 consecutive patients (median follow-up 59 months), 30 received full standard administered mean activity of 25.5 GBq and 35 received reduced administered mean activity of 17.8 GBq. No occurrence of grade 3 or 4 hematologic toxicity or MDS/AL was seen. The median OS was not reached by the standard dose cohort and was 63.8 months for the reduced dose cohort with an overall DCR of 87%. This update once again demonstrates the safety and efficacy of ¹⁷⁷Lu-DOTATATE PRRT, and furthermore, supports personalized dose scheduling based on patient baseline factors.²⁹

Baum et al.³⁰ presented their large single-center experience of ⁹⁰Y and/or ¹⁷⁷Lu PRRT for GEPNETs from Bad Berka. The minimum duration of follow-up was 2 years with a median >5 years. Of the 1048 patients treated, 21 (2%) developed MDS and 2 (<1%) AL. The median OS for the entire cohort was 51 months with a median OS of 19 months for those who developed MDS. Improved survival was noted in patients <40 years, treated with a combination of both ⁹⁰Y and ¹⁷⁷Lu PRRT with grade 1 tumors of intestinal origin.³⁰

Prasad et al. (personal communication 2016 ENETS) also report encouraging findings of long-term safety for PRRT for GEPNET patients in Berlin. None of their 150 patients, treated with ¹⁷⁷Lu PRRT monotherapy at a standard administered activity of 7.4 GBq, has developed MDS or AL over the median follow-up period of 3–4 years. Furthermore, they have noted sustained durable response rates.

The efficacy and safety of PRRT monotherapy for NETs have been well established and now consolidated with robust long-term data spanning 15 years of experience. Despite this, given the nature of NET, there is an inherent concern regarding relapsed disease, and the introduction of novel agents has led researchers to explore their efficacy in combination with PRRT in an effort to improve and sustain response rates.

Lutetium-177-DOTATATE in combination with radiosensitizing chemotherapy

van Essen et al. in Rotterdam³¹ were the first to publish short-term hematotoxicity data in their interim report after a 1-year follow-up of seven patients treated with ¹⁷⁷Lu-DOTATATE at an administered activity of 7.4 GBq plus capecitabine 1650 mg/m² daily for 14 days. The patients were all men, with a median age of 62, without prior exposure to either chemotherapy or PRRT. Six patients completed four cycles of therapy and one patient two cycles. Two of the seven developed grade 3 toxicity; one of hemoglobin and one of platelets. The patients who developed grade 3 anemia made a spontaneous recovery within 2 weeks but went on to develop progressive disease. The patient experiencing thrombocytopenia underwent a break from therapy and resumed at a lower dose without any recurrence of toxicity. No patient developed grade 4 hematotoxicity or any significant leukopenia. Thus, addition of capecitabine was noted to be “feasible and safe” with no clear associations between therapy, clinical features and development of toxicity documented in this preliminary study.³¹

Claringbold et al.⁵ treated 33 patients in their Phase II Australian clinical study of standard 7.8 GBq ¹⁷⁷Lu-DOTATATE every 8 weeks in combination with capecitabine 1650 mg/m² daily for 14 days. The majority of patients completed the intended four cycles of therapy. Only one patient developed grade 3 thrombocytopenia, which was self-limiting and spontaneously resolved. No neutropenia was documented. Furthermore, bone marrow activity was not evident on whole body single positron emission computed tomography (SPECT/CT) whole-body post-therapy 24-hour images, consistent with the reported comparable activities in blood and bone marrow and the absence of ¹⁷⁷Lu-DOTATATE binding to bone marrow stem cells. All interruptions of capecitabine were due to idiosyncratic angina. During the 6-month follow-up period, no case of MDS/AL was seen. Thus, the addition of capecitabine to conventional PRRT did not increase the modest reversible short-term toxicity that had been noted with ¹⁷⁷Lu-DOTATATE monotherapy in historic PRRT cohorts at the same institution.⁵

When the efficacy of capecitabine/temozolomide chemotherapy in pancreatic NETs was reported by Strosberg et al.³² Claringbold et al. went on to conduct a Phase I–II study of ¹⁷⁷Lu-DOTATATE combined with capecitabine and temozolomide.⁶ Temozolomide was administered on days 10–14 with the phase one component of the study identifying 200 mg/m² as the optimal dose. Thirty-four patients completed four cycles of treatment at 8-week intervals. The addition of temozolomide to the standard ¹⁷⁷Lu-DOTATATE-capecitabine regimen was associated with a higher incidence of low-grade reversible thrombocytopenia (29% grade 1–2) and neutropenia (18% grade 1–2 and 6% grade 3). Although of a lower grade, thrombocytopenia was the commonest reason for dose reduction, and prior chemotherapy was once again identified as a risk factor.⁶ This study clearly demonstrated that the addition of an alkylating agent to PRRT did not significantly increase the incidence of short-term toxicity over a follow-up period of 24 months, as long as any early toxicity was appropriately managed. Furthermore, the DCR of 94% suggested enhanced efficacy of this approach, especially the achievement of high ORR; complete response (CR) of 16%, and partial response (PR) of 41%, in comparison to PRRT monotherapy.

Claringbold et al. further documented the safety of combination therapy in 2014 when they retrospectively reviewed the long-term outcomes of both cohorts of ¹⁷⁷Lu-DOTATATE combined chemotherapy patients. Of the 65 patients analyzed (28 treated with ¹⁷⁷Lu-DOTATATE+capecitabine, 37 treated with ¹⁷⁷Lu-DOTATATE+capecitabine+temozolomide), short-term toxicity over 6 months was reversible and self-limiting. Only one patient (3.5%) in the PRRT capecitabine cohort developed grade 3 toxicity of neutrophils. A higher incidence of grade 1/2 thrombocytopenia was noted in the PRRT capecitabine and temozolomide group (78.5% vs. 60.5%). Long-term follow-up (median of 60 months for PRRT capecitabine and 36 months for PRRT capecitabine+temozolomide) revealed an increased incidence of grade 3/4 toxicity in the temozolomide group (six patients vs. two patients), manifesting as grade 3 anemia (four patients), grade 3 neutropenia (one patient), and grade 4 thrombocytopenia (one patient). Two patients in the temozolomide cohort developed MDS (5.4%), each of whom was aged >65 years, had complex cytogenetic karyotypes, and had been heavily pretreated (chemotherapy, pelvic irradiation, and sunitinib). Thus, any causal relationship between the addition of temozolomide and development of MDS in these patients is conjectural. The most significant factor predicting myelotoxicity in this study was the presence and number of prior therapies, with heavily pretreated patients at an increased risk of both short- and long-term toxicity.³³

More recently, the Australian group of Claringbold and Turner⁷ has gone on to explore in a Phase I study, the combination of PRRT with everolimus, a potent inhibitor of the mammalian target of rapamycin (mTOR), which is an approved agent with efficacy in treatment of NETs. The NETTLE study recruited 16 heavily pretreated patients in three cohorts with escalating everolimus daily doses of 5, 7.5, and 10 mg in combination with standard 7.8 GBq ¹⁷⁷Lu-DOTATATE cycles. Everolimus therapy was combined with the first three cycles of PRRT and omitted from the fourth and final cycle. Over the short-term follow-up period of 6 months, three patients developed grade 3 toxicity (two of platelets and one of neutrophils), none requiring treatment. The most significant associated factors were everolimus daily dosing at 10 mg and a history of chemotherapy. All patients in the 10 mg everolimus/PRRT cohort demonstrated dose-limiting toxicity requiring dose reduction. The MTD of everolimus was found to be 7.5 mg/day in combination with PRRT, and some patients required a further daily dose reduction to 5 mg everolimus. Patients who maintained adequate counts (≥grade 2 NCTCAE v4 criteria for hemoglobin, platelets, and neutrophils) at week 12 of therapy were unlikely to experience any further deterioration. Similar to findings in other reported PRRT studies, these “higher risk” patients could usually be identified early in their treatment course. An ORR of up to 80% in pancreatic NET was achieved which is an order of magnitude greater than that reported for everolimus alone (8% versus 80%).⁷

Everolimus monotherapy as salvage following PRRT

Kamp et al. from Rotterdam³⁴ retrospectively reviewed outcomes of everolimus monotherapy at daily doses of 5 or 10 mg as salvage in patients who had progressed following previous ¹⁷⁷Lu-DOTATATE PRRT. Of the 24 patients treated (median age 60 years, median follow-up 11 months), 3 developed grade 3/4 toxicity (2 of platelets and 1 of WBC) with no incidence of MDS. Coupled with a median PFS of 13.1 months, the study concluded that the safety profile of everolimus is not influenced by previous PRRT.³⁴

These findings were in direct contrast to those of Panzuto et al. from Milan,³⁵ who managed progressive disease in 164 heavily pretreated patients (median age 63 years, median follow-up period of 12 months) with everolimus 10 mg and 5 patients with 5 mg per day. One hundred forty-seven patients received both everolimus and somatostatin analog therapy. Twenty-one of the 169 patients (12.4%) experienced grade 3/4 myelotoxicity (13 of platelets and 9 of hemoglobin. No data on WBC toxicity were reported) with a strong correlation to prior chemotherapy or PRRT (as monotherapy or in combination, p < 0.01). Everolimus therapy was discontinued in 103 of 169 patients, predominantly because of disease progression (50.3%), but a small subset (8.3%) due to toxicity. The authors thus urged caution when using everolimus in heavily pretreated patients, suggesting that it should be used earlier in the treatment course of NETs.³⁵

Discussion

The advent of PRRT has revolutionized the management of patients suffering with NETs. Within the current therapeutic landscape, all major international NET guidelines include PRRT in their algorithms, however, the optimal timing of implementation remains contentious, and sequencing and combination of therapies are controversial.

There is clear evidence that myelotoxicity does occur with PRRT. Appropriate timing of therapy, careful patient selection, dose optimization, and adequate monitoring are mandated, to minimize the risk of significant long-term sequelae, including MDS and AL.

The agent of choice for PRRT, either alone or in combination, is ¹⁷⁷Lu-DOTATATE, owing to its high affinity for the SSR2 receptor, and limited range of emitted β radiation. The concomitant low abundance γ emission of an optimal energy for quantifiable γ camera imaging provides an opportunity for individualized dosimetry and monitoring of metabolic therapeutic response after each cycle of ¹⁷⁷Lu-DOTATATE PRRT.

Lutetium-177-PRRT allows optimization of tumor-targeted dose with reduced critical organ toxicity, especially with respect to myeloid tissue. Short-term toxicity of ¹⁷⁷Lu-DOTATATE PRRT-based therapy commonly presents as reversible, limited grade cytopenias preferentially affecting platelets, WBC, and hemoglobin. The timing of platelet toxicity is suggestive of megakaryocyte (MK) sensitivity to, and modulation of, endomitosis/endoreduplication, as a result of myeloid exposure to radiation. Given that a single MK will bud to produce numerous platelets, the platelet nadir reached is more a reflection of impaired production. However, the sequential processes involved in MK maturation include numerous signaling molecules, microRNAs, transcriptional factors, and their downstream target genes.³⁶ The crossfire and innocent bystander effects PRRT clearly potentiate damage to the marrow microenvironment on both a cellular and genetic level. This is analogous with the finding that manifest myelotoxicity arising early in the treatment course is a poor prognostic feature. It is associated with an increased risk of developing long-term toxicity and necessitates dose modification, or therapy cessation in the setting of combined treatments. Long-term toxicity in the form of MDS/AL is of low incidence and is delayed, occurring 5–10 years post-therapy. This observation makes interpretation of possible causal factors difficult, being confounded by toxicities attributable to prior treatments, other comorbidities, medications, and the aging process.

The incidence and pattern of toxicity are not greatly affected by the addition of alkylating agents or radiosensitizing chemotherapy and only modestly increased with concomitant use of mTOR inhibitors with ¹⁷⁷Lu-DOTATATE PRRT. However, more recent studies have clearly identified that salvage therapy of heavily pretreated patients with PRRT monotherapy appears to result in both an increased incidence and significance of myelotoxicity.

It is evident from the studies reviewed that the risk factors for development of both short/long-term toxicity include the number of prior therapies; exposure to chemotherapy with alkylating agents, radiation-based therapy, and PRRT; age >65 years; impaired renal function; depleted myeloid reserve (manifesting as baseline cytopenias/early development of significant grade toxicity, either due to bone marrow involvement or resulting from prior therapies); and poor performance status. These adverse features, if present, warrant consideration of alternate therapies or substantial reduction of administered activity of ¹⁷⁷Lu-DOTATATE PRRT.

Of the risk factors, prior exposure to chemotherapy, especially alkylating agents, correlates strongly with the development of long-term toxicity, including MDS and subsequent AL. Therapy-related MDS has a distinct presentation with abnormalities of chromosomes 5 and 7, complex karyotypes (>3 cytogenetic abnormalities), and clinically a very rapid transformation to AL leading to limited survival. These classical features of therapy-related MDS have only been documented in a small number of the studies reviewed and warrant further detailed consideration in future studies, with the possible incorporation of a standard bone marrow assessment, including aspirate morphology and karyotyping studies at baseline as a risk-stratifying tool. Such a strategy would also provide a robust measure of mutation acquisition and in future may aid in profiling, yet-to-be-determined “high-risk,” abnormalities that may be associated with PRRT-based therapies.

The association of prior therapy to MDS/AL is suggestive of cumulative toxicity from PRRT. Denoyer et al. objectively demonstrated this in NET patients treated with ¹⁷⁷Lu-DOTATATE in their 2015 study, utilizing serial measurements of phosphorylated histone variant H2AX (γ-H2AX) by immunofluorescence as a surrogate marker for external-beam ionizing radiation-induced DNA double-strand breaks (DSBs). Similar trends were noted in all 11 patients studied, with notably increased γ-H2AX foci in circulating lymphocytes relative to pretherapy values, with peak toxicity noted at 0.5 to 4 hours of post-therapy, subsequently decreasing by 24 to 72 hours. Foci number directly correlated with absorbed dose to tumor, bone marrow, and the extent of peripheral blood lymphocyte reduction. However, the predicted level of γ-H2AX foci was substantially less than the experimental values, indicating that the radiation dose only partially induces the DSBs detected. Furthermore, the number of residual foci at 24 to 72 hours postdose was higher than that predicted, possibly due to inefficient DNA repair and/or mobilization of damaged lymphocytes from high-uptake tissues (spleen, bone marrow, and tumor). Despite the variability in patients included, those who had prior exposure to 5-fluorouracil had a significantly increased level of γ-H2AX foci, suggesting increased sensitivity of lymphocytes to DNA damage.³⁷

Eberlein et al. recently published their calibration of the γ-H2AX DSB focus assay for internal radiation exposure, thus demonstrating its potential application in PRRT dosimetry.³⁸ In their 2015 article, they studied radiation-induced induction of colocalizing γ-H2AX and 53BP1 foci as a surrogate for DSB. After developing a calibration curve by correlating foci per cell values with in vitro absorbed doses to blood from ¹³¹I and ¹⁷⁷Lu, they used 55 normal blood samples (from three healthy individuals) spiked with radioactive solutions of various concentrations of isotonic NaCl-diluted radionuclides of known activity. Lymphocytes isolated were ethanol fixed and underwent two-color immunofluorescence staining for frequencies of the colocalizing γ-H2AX and 53BP1 foci/nucleus. This methodology allowed calculation of absorbed dose rates to the blood per nuclear disintegration occurring in 1 mL of blood and was calculated for both isotopes. The investigators were able to confirm a linear relationship between the number of DSB-marking foci/nucleus and the absorbed dose to blood from both ¹³¹I and ¹⁷⁷Lu. Although their approach of ethanol fixation and dual-color flow cytometry is impractical for routine use, the principle of the assay is clearly adaptable for use with conventional peripheral whole blood flow cytometric analysis, once the appropriate antibodies and fluorochromes are developed. This marks a paradigm shift to potential on-treatment dynamic dosimetry analysis, which would clearly lend itself to an individualized risk-adapted PRRT approach.

Although individualized dosimetry methods for PRRT have evolved over the last decade, as of yet, a standardized approach has not been established.³⁹ Ljungberg et al. in their 2015 publication outlined the current EANM/MIRD guidelines for use of quantitative ¹⁷⁷Lu SPECT for applied dosimetry. Given the biexponential activity of ¹⁷⁷Lu, multiple SPECT/CT images at defined time points following dose administration are required to assess whole-body activities. Regarding myelotoxicity, imaging at time 0 and 24 hours following infusion, as outlined, would not capture sufficient information regarding the fast phase decay that contributes to acute toxicity. To fully evaluate this early activity, at least six SPECT/CT imaging studies would be required between time 0 and 24 hours, which is clearly not feasible both in terms of resources and patient convenience. Thus, there is a clear imperative to not only standardize a method for whole-body dosimetry but to also validate a convenient, cost-effective, novel methodology, once again highlighting the importance of methods such as DSB analysis.

Regarding the longer term risks of MDS/AL, in 2008, Yokoyama and Cleary described the relationship between the tumor suppressor menin (a product of the MEN1 gene, located on 11q13 and associated with GEPNETs) and the mixed-lineage leukemia histone methyltransferase complex (MLL-HMT, located on 11q23).⁴⁰ MLL is required to promote progenitor expansion and stem cell self-renewal in the hematopoietic lineage. In a subset of AL, chromosomal aberrations resulting from DNA damage generate chimeric MLL oncoproteins resulting in aberrant transcription factor expression and leukemogenesis. The article clearly outlined the leukemogenic role of menin as a molecular adaptor for MLL-HMT interaction with lens epithelium-derived growth factor (LEDGF), a chromatin-associated protein implicated in autoimmunity, HIV-1, and leukemia. This article was one of the first to establish the discordant function of menin as a tumor suppressor in the endocrine lineage and as a promoter of oncogenesis in the hematopoietic lineage.

In addition, Li et al.⁴¹ further elaborated upon the complex role of the MLL family of proteins in their 2016 article. The severely compromised methyltransferase activity of MLL1 was shown to be significantly increased by its interaction with minimized human RBBP5-ASH2L heterodimer in a two-step mechanism, not only providing an insight into the assembly and activity regulation of the MLL family of methyltransferases but also raising the possibility of a universal “switch” for regulation of histone methyltransferases and their complex interaction with other proteins.

Although clinically, lymphocyte toxicity alone seldom requires intervention, the ability to measure DNA damage as described by Denoyer et al., combined with the genetic link between MEN1 and MLL, raises a fundamental issue in the management of GEPNET patients. It may be that these patients are inherently at risk of leukemogenesis and the common relapsing/refractory course exposing them to exponential DNA damage eventually results in an imbalance in favor of the hematopoietic oncogenic role of menin through its molecular cofactor activity with MLL-HCT. Thus, future treatment protocols should aim to incorporate specialized tissue studies (tumor and bone marrow) such as fluorescence in situ hybridization (focusing on 11q13/11q23) and cytogenetics in combination with methods for monitoring DNA damage as described.

The increased incidence of MDS/AL in relation to prior therapies raises another interesting observation regarding PRRT prescribing practices. This review clearly demonstrates the adverse outcomes inherent in an approach that relegates PRRT to a last resort salvage treatment option.

Given the marked improvement in DCRs since the introduction of PRRT, it is now time to acknowledge that this therapy, if available and considered appropriate, should be ideally administered to GEPNET patients first line, or on progression following long-acting somatostatin analogs (SSA). This would not only serve to mitigate significant cumulative myelotoxicity but may be expected to translate into further improvements in response. This last point is of particular importance given the efficacy that is demonstrated with PRRT combination therapies, and taking into account the increased incidence of toxicity noted with salvage treatment strategies.

Novel agents targeting receptors involved in NET biology are now becoming more widely available. PRRT combination with agents, such as everolimus, has already demonstrated short-term safety and efficacy in early-phase clinical studies. New combinations of PRRT with immune checkpoint inhibitors are being contemplated to improve outcomes compared with PRRT alone or PRRT in combination with chemotherapy.

With improved diagnostic imaging and an expanding repertoire of novel agents becoming available, a relative tsunami of trials studying PRRT-based therapies for NETs is expected. The limited data available on novel therapies suggest that to improve tolerability, mitigate toxicity, and improve response outcomes, they are best administered in combination with PRRT and not as salvage following PRRT. Furthermore, early therapy with PRRT-containing regimens for GEPNETs improves outcomes and minimizes toxicity.

The evidence base for combination ¹⁷⁷Lu-DOTATATE-capecitabine/temozolomide is currently being formalized in a multicenter Australian Phase II randomized controlled trial, CONTROL-NETS (ACTRN12615000909527), under the auspices of the Australasian Gastrointestinal Trials Group and is designed to define toxicity and efficacy while addressing the unmet need for a standard evidence-based, accepted PRRT combination protocol with respect to sequencing and chemotherapy/biologic agents. Meanwhile, the new diagnostic gold standard for NETS, ⁶⁸Ga-DOTATATE PET/CT, is becoming widely available and the theranostic approach of ⁶⁸Ga/¹⁷⁷Lu-DOTATATE to PRRT will play a major part in improving NET patient therapy outcomes by improving diagnosis, patient selection, and tumor response monitoring.

Footnotes

Disclosure Statement

No competing financial interests exist.

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