3. DISEÑO METODOLÓGICO DE INVESTIGACIÓN
3.5. Material experimental
On the whole any beneficial effect o f MDR modulation in adult patients has been
disappointing. The most encouraging report is of three responses in eight patients (seven
with multiple myeloma and one with lymphoma) who progressed on treatment with
VAD (continuous infiision vincristine and doxorubicin with oral dexamethasone), and
then went on to receive VAD with the addition of a continuous infusion of verapamil
(Dalton et al 1989).
i Verapamil
Verapamil has been widely evaluated as a resistance modifier in adults with a range of
malignant diseases. In most cases high dose racemic verapamil has been given by
continuous intravenous infusion. There are however a few reports using oral racemic
verapamil or D-verapamil.
Both verapamil and its principle metabolite norverapamil are active in modulating MDR
in vitro (Merry et al 1989), however norverapamil has fewer cardiovascular effects
(Hamann et al 1984). In clinical practice concentrations of up to 3000 ng/ml are
possible (Dalton et al 1989), though more realistically, concentrations o f 200 - 500
ng/ml can be maintained with little toxicity (Benson et al 1985). However serum levels
do not accurately reflect tissue or tumour levels which tend to be higher (Hamann et al
1984).
The first reported phase I study using continuous infusional verapamil to modulate
MDR was reported in 1985 (Benson et al 1985), using a loading dose of verapamil (0.02 - 0.1 mg/kg) followed by a maintenance infusion (0.036 - 0.18 mg/kg/hr) for 132 houra.
for five days) to 17 adults with a variety o f advanced relapsed malignant diseases. The
observed cardiovascular toxicity consisted of first degree heart block, junctional
rhythms and non specific T wave changes. Other toxicity included haematological
(neutropenia and thrombocytopenia), and neurological (peripheral neuropathy and
myalgia). The steady state verapamil concentrations achieved were between 200 - 400
ng/ml for a continuous infusional dose o f 0.12 mg/kg/hr.
An alternative approach (Ozols et al 1987) was to treat each patient (all had advanced
ovarian cancer) with a continuous infusion of verapamil in a dose escalating fashion
until hypotension or heart block developed. This dose was then maintained for 72 hours,
with a 24 hour infusion of adriamycin given on the second day of the verapamil
infusion. All patients were electively treated on the intensive care unit, two of the eight patients treated developed atropine responsive complete heart block, and four patients
developed congestive heart failure. Delivery of these doses of verapamil resulted in
serum verapamil concentrations of up to 2767 ng/ml (about 6 pM), which is the level
required to reverse resistance in vitro (Tsuruo et al 1982, Twentyman et al 1986b).
There were no objective responses to this regimen, though two patients had stabilisation
of their disease.
Cardiovascular toxicity is the most commonly reported dose limiting factor in trials
attempting to escalate the administered dose of verapamil (Ozols et al 1987, Miller et al
1991, Salmon et al 1991). Other side effects commonly attributed to verapamil rarely
influence the decision to reduce or to stop the infusion.
et al 1989, Dalton et al 1989). For example Cairo (1989) used a continuous infusion of
verapamil (0.005 mg/kg/h) and achieved steady state concentrations o f 450 ng/ml, 8/11
patients achieved a PR (a mixed group of paediatric tumours). Dalton (1989) used a
continuous infusion of verapamil (0.15 mg/kg) in patients v^th multiple myeloma and
achieved a steady state concentration of 200 - 300 ng/ml in two patients who achieved a
PR, and 1342 ng/ml in one patient who attained a temporary CR, though this patient
received a higher dose of verapamil.
It may be that it is tissue or tumour concentration which is important, or because the
amount of verapamil required to modulate MDR in vivo is less than that shown to be
necessary in vitro.
Continuous low dose racaemic verapamil has been used in patients with anthracycline
resistant breast cancer (Belpomme et al 1994). 99 patients were treated, half with vindesine and 5 FU alone and half v^th additional verapamil. Verapamil was given
orally at a dose of 120 mg twice a day for 28 days, this resulted in few side effects when
compared with patients not receiving it. The response rate in the chemotherapy alone
arm was 5/47 compared v\dth 14/52 in those also receiving verapamil. This suggests a
benefit from continuous low dose verapamil, though the mechanism is unclear.
Oral verapamil has been used in adults to try and provide an effective regimen which is
less toxic than intravenous verapamil. There is evidence (Hamann et al 1984) that when
verapamil is administered orally the hepatic first pass effect differs from that seen with
intravenous administration, resulting in lower concentrations of the more cardiotoxic L-
isomer and higher concentrations of the D-isomer.
Two o f the published studies were randomised, one in small cell lung cancer
et al 1993), both used the same dose of verapamil (480 mg/day in divided doses) for
five or three days respectively. Neither study had significant problem with
cardiovascular toxicity, though the NSCLC study had an increased rate of neurological
problems (principally constipation and peripheral neuropathy) in the verapamil arm.
The SCLC study (Milroy et al 1993) randomised 226 patients and found no difference
in either response rate or overall survival. The study in NSCLC (Millward et al 1993)
recruited 72 patients, 66 of these were evaluable for response. This study showed a
significant improvement in overall response rate (41% versus 18%), and some
improvement in survival (all patients were dead at 96 weeks in the 'no verapamil' arm,
whilst two were still alive at six months in the verapamil arm).
D-verapamil has been used in an attempt to overcome the cardiovascular side effects
seen with the racemic agent following the observation that both optical isomers of
verapamil are equally effective in vitro as MDR modulators (Plumb et al 1990, Hollt et
al 1992, Woodcock et al 1993). These studies (Bissett et al 1991, Scheithauer et al
1993) suggest that whilst D-verapamil is less cardiotoxic than the racemic mixture a
significant number of patients develop cardiovascular side effects until the dose is
dropped to 800 mg/day or less (resulting in a serum level of about 1000 ng/ml) higher
serum concentrations have been made tolerable however by the addition of
dexamethasone. There is also the observation that when given with an anthracycline, the
resulting myelotoxicity was greater than expected, implying that a pharmacokinetic
interaction occurs between the cytotoxic agent and D - verapamil (and not with racemic
verapamil). Out of ten patients treated on one of these studies (Bissett et al 1991) there
A more recent study (Bates et al 1994) treated 66 patients (relapsed lymphoma) with a
regimen consisting of chemotherapy (etoposide, prednisolone, vincristine
cyclophosphamide and doxorubicin) and D-verapamil (dose escalating to a maximum of
1200 mg/day). They found cardiovascular toxicity to be dose limiting though reduced if
D-verapamil was given 4 hourly rather than 6 hourly. Twelve of the 62 evaluable
patients had a response to this, including two complete responses. MDR 1 expression
(PCR method) was quantified in some of the patients pre, peri and post treatment. In
some (7/15) there was an increase in MDR 1 following chemotherapy alone, these
patients appeared to do better (in terms of response) when D-verapamil was added to
chemotherapy.
Whilst a wide range of modulating agents are being investigated in the laboratory, the
number of agents used in clinical trials remains limited. Other calcium channel blockers
have been evaluated, including diltiazem (Bessho et al 1985), nifedipine (Philip et al
1992) and bepridil (Linn et al 1994), however use of these is also restricted by dose
limiting cardiovascular toxicity.
ii Cyclosporin
The modulating agent generating most interest recently is cyclosporin, which has been
used in acute leukaemia (Sonneveld et al 1990, List et al 1993), multiple myeloma
(Sonneveld et al 1992), and in a collection of relapsed and resistant malignancies
(Yahanda et al 1992, Bartlett et al 1994). The regimens used generally consist o f a
loading dose of up to 6 mg/kg over two hours followed by a continuous infusion of up
to 18 mg/kg/day for at least 60 hours. In a series of 42 patients with acute myeloid
myeloma (Sonneveld et al 1992) an overall 48% response rate included a more
favourable subgroup of patients vs^ho expressed MDR 1 and had a 58% response rate.
List (1993) treated 42 patients with poor risk AML with cytarabine and daunorubicin.
All of these patients received a CSA infusion for 72 hours (i.e. coincident Avith the
daunorubicin infusion). The dose of CSA was escalated from a loading dose o f 1.4 to 6
mg/kg and a continuous infusion dose of 1.5 to 20 mg/kg/day. Steady state CSA
concentrations of greater than 1500 ng/ml were achieved in all patients receiving more
than 16 mg/kg/day continuous infusion. 26 patients achieved a CR or restored chronic
phase, 3 a PR giving an overall response rate of 69%.
Patients with VAD refractory multiple myeloma were treated (Sonneveld et al 1992)
with identical chemotherapy (VAD) except for the addition o f a continuous infusion of
CSA ( 5 - 1 0 mg/kg/day). All patients who received 7.5 mg/kg/day or more achieved
serum CSA concentrations of at least 1000 ng/ml. There were 17 responders, and in 6 of
these no MDR 1 positive (C219 by IHC) myeloma cells could be detected.
Concern about the cyclosporin related alteration in etoposide pharmacokinetics has lead
to the suggestion that etoposide concentrations need to be measured and the dose
adjusted when given concurrently with cyclosporin (Yahanda et al 1992). This would
allow a meaningful comparison to be made between the response seen with etoposide
given alone, and the response seen with subsequent courses of treatment where
etoposide is given with cyclosporin. In this way the additional benefit (in terms of
disease response) can be assessed and excess toxicity avoided.
PSC 833 has been used in phase I studies in combination with etoposide, taxol and
VAD (vincristine, doxorubicin and dexamethasone) in adult patients with solid tumours
Etoposide or taxol alone were given to patients with resistant tumours (Lum et al 1994,
Fisher et al 1994), if there was no response to this oral PSC 833 alone was given (to
assess response and pharmacokinetics). Further courses consisted of PSC 833 and
cytotoxic (with assessment of response, adverse events and PSC 833 pharmacokinetics).
As seen by other groups the dose limiting toxicity of PSC 833 was cerebellar with
reduced proprioception and ataxia, this effect was most marked 2 hours after each oral
dose and in some cases persisted for weeks. Etoposide doses had to be reduced by more
than 50% when given with PSC 833 to achieve the expected degree of
myelosuppression. In this (and other) studies a marked difference in the bioavailiblity of
different oral preparations was seen.
Thirty five patients (relapsed solid tumours) have been treated at Cambridge
(unpublished meeting report 1994) with intravenous etoposide and PSC 833. Significant
increases in etoposide AUC were demonstrated when given with PSC 833 up to a PSC
dose o f 4 mg/kg/day, above this there was no further change. Severe neurotoxicity was
observed in two patients with many more experiencing minor neurological events
(paresthesia or dizziness), hyperbilirubinaemia, nausea and vomiting.
In VAD refractory multiple myeloma patients (Sonneveld et al 1994) given VAD with
PSC 833, all cytotoxic drugs had to be dose reduced by 50% - 75% (following the first
course) on the basis of toxicity and doxorubicin concentrations. Adverse events were
similar to those in other studies except for a higher incidence of constipation (attributed